Publications

Long-term genetic studies of wild populations are very scarce, but are essential for connecting ecological and population genetics models, and for understanding the dynamics of biodiversity. We present a study of a wild wheat population sampled over a 36-year period at high spatial resolution. We genotyped 832 individuals from regular sampling along transects during the course of the experiment. Genotypes were clustered into ecological microhabitats over scales of tens of metres, and this clustering was remarkably stable over the 36 generations of the study. Simulations show that it is difficult to determine whether this spatial and temporal stability reflects extremely limited dispersal or fine-scale local adaptation to ecological parameters. Using a common-garden experiment, we showed that the genotypes found in distinct microhabitats differ phenotypically. Our results provide a rare insight into the population genetics of a natural population over a long monitoring period.

This open access book covers a century of research on wheat genetics and evolution, starting with the discovery in 1918 of the accurate number of chromosomes in wheat. We re-evaluate classical studies that are pillars of the current knowledge considering recent genomic data in the wheat group comprising 31 species from the genera Amblyopyrum, Aegilops, Triticum, and other more distant relatives. For these species, we describe morphology, ecogeographical distribution, phylogeny as well as cytogenetic and genomic features. For crops, we also address evolution under human selection, namely pre-domestication cultivation and domestication. We re-examine the genetic and archeological evidence of where, when, and how domestication occurred. We discuss unique aspects of genome evolution and maintenance under polyploidization, in natural and synthetic allopolyploids of the wheat group. Finally, we propose some thoughts on the future prospects of wheat improvement. As such, it can be ofgreat interest to wheat researchers and breeders as well as to plant scientists and students interested in plant genetics, evolution, domestication, and polyploidy.

Bread wheat (Triticum aestivum, genome BBAADD) is a young hexaploid species formed only 8,500-9,000 years ago through hybridization between a domesticated free-Threshing tetraploid progenitor, genome BBAA, and Aegilops tauschii, the diploid donor of the D subgenome. Very soon after its formation, it spread globally from its cradle in the fertile crescent into new habitats and climates, to become a staple food of humanity. This extraordinary global expansion was probably enabled by allopolyploidy that accelerated genetic novelty through the acquisition of new traits, new intergenomic interactions, and buffering of mutations, and by the attractiveness of bread wheat's large, tasty, and nutritious grain with high baking quality. New genome sequences suggest that the elusive donor of the B subgenome is a distinct (unknown or extinct) species rather than a mosaic genome. We discuss the origin of the diploid and tetraploid progenitors of bread wheat and the conflicting genetic and archaeological evidence on where it was formed and which species was its free-Threshing tetraploid progenitor. Wheat experienced many environmental changes throughout its evolution, therefore, while it might adapt to current climatic changes, efforts are needed to better use and conserve the vast gene pool of wheat biodiversity on which our food security depends.

Recombination between homeologous chromosomes, also known as homeologous exchange (HE), plays a significant role in shaping genome structure and gene expression in interspecific hybrids and allopolyploids of several plant species. However, the molecular mechanisms that govern HEs are not well understood. Here, we studied HE events in the progeny of a nascent allotetraploid (genome AADD) derived from two diploid progenitors of hexa- ploid bread wheat using cytological and whole-genome sequence analyses. In total, 37 HEs were identified and HE junctions were mapped precisely. HEs exhibit typical patterns of homologous re- combination hotspots, being biased toward low -copy, subtelo- meric regions of chromosome arms and showing association with known recombination hotspot motifs. But, strikingly, while homologous recombination preferentially takes place upstream and downstream of coding regions, HEs are highly enriched within gene bodies, giving rise to novel recombinant transcripts, which in turn are predicted to generate new protein fusion variants. To test whether this is a widespread phenomenon, a dataset of high - resolution HE junctions was analyzed for allopolyploid Brassica , rice, Arabidopsis suecica , banana, and peanut. Intragenic recombi- nation and formation of chimeric genes was detected in HEs of all species and was prominent in most of them. HE thus provides a mechanism for evolutionary novelty in transcript and protein se- quences in nascent allopolyploids.

The origin and evolution of the wheat group (the genera Aegilops, Amblyopyrum, and Triticum) in the wild and under cultivation is reviewed. The diploid species diverged from a common ancestor, about 24 million years ago, presumably in the marginal Mediterranean region of Southwest Asia. Speciation resulted in distinct genomes, specialized dispersal unit, and unique eco-geographical environments that are described here. In contrast to the diploids, the allopolyploid species have undergone convergent evolution, exhibiting a wider variation, larger distribution areas and a broader range of ecological conditions. Recent molecular studies show reprograming of gene expression in the allopolyploids and the interactions between genomes. The process of allopolyploidization generates a genetic shock that triggers a variety of cardinal genetic and epigenetic changes leading to cytological and genetic diploidization, enabling various gene dosage effects to take place and providing tolerance to hybridization and introgression.

Subgenome integrity in bread wheat (Triticum aestivum; BBAADD) makes possible the extraction of its BBAA component to restitute a novel plant type. The availability of such a ploidy-reversed wheat (extracted tetraploid wheat [ETW]) provides a unique opportunity to address whether and to what extent the BBAA component of bread wheat has been modified in phenotype, karyotype, and gene expression during its evolutionary history at the allohexaploid level. We report here that ETW was anomalous in multiple phenotypic traits but maintained a stable karyotype. Microarray-based transcriptome profiling identified a large number of differentially expressed genes between ETW and natural tetraploid wheat (Triticum turgidum), and the ETW-downregulated genes were enriched for distinct Gene Ontology categories. Quantitative RT-PCR analysis showed that gene expression differences between ETW and a set of diverse durum wheat (T. turgidum subsp durum) cultivars were distinct from those characterizing tetraploid cultivars per se. Pyrosequencing revealed that the expression alterations may occur to either only one or both of the B and A homoeolog transcripts in ETW. A majority of the genes showed additive expression in a resynthesized allohexaploid wheat. Analysis of a synthetic allohexaploid wheat and diverse bread wheat cultivars revealed the rapid occurrence of expression changes to the BBAA subgenomes subsequent to allohexaploidization and their evolutionary persistence.

Wild emmer wheat, Triticum turgidum subsp. dicoccoides, (2n = 4× = 28; genome BBAA), the progenitor of domesticated wheat, is genetically closely related to durum and common wheat. This wild taxon has characteristics that would be valuable if transferred to domesticated wheat. A series of chromosome-arm substitution lines (CASLs) of wild emmer wheat were produced in the background of an Israeli common wheat cultivar. These CASLs were evaluated in a pot experiment and in field trials in Israel and California for their grain yield (GY) and its components and for grain protein percentage. In addition, the extent of genetic interactions (epistatic effects) between "wild" and "domesticated" alleles, within and between homoeologous groups 1 and 7 as expressed in grain and protein yields and other quantitative traits, were determined. The research has shown that wild emmer wheat harbors genetic variability for quantitative traits and that the "wild" genes interact among themselves in a non-additive way in the common genetic background. Several chromosome arms improve GY and protein percentage in common wheat, but their effects will be presumably enhanced when combination of genes from several "wild" arms are integrated into a single "domesticated" genotype. Hence, the interaction between these genes and those in the recipient common wheat must be accounted for when higher yield or protein content is desired. The results of this study indicate on the potential of this material for breeding and genetic analysis, and support the idea of pyramiding genes from a wild species.

Durum wheat appears in the archaeological record, very sporadically, ~7000 years before present, but becomes the dominant tetraploid wheat in the Levant and in the Mediterranean basin ~2500 years ago. Here, we discuss the archeological insights on durum wheat evolution and we focus on the analysis of the genomic changes that are correlated with the process of domestication and evolution of modern durum by comparing four genetic groups: wild emmer, domestic emmer, durum landraces and modern durum varieties. Changes in gene expression and copy number variation of genes and transposons were analyzed in the genetic groups. Genes were clustered based on their pattern of change during Durum evolution, e.g. gradual increase, or decrease, or increase at the onset of domestication and plateauing later on. There were not many genes that changed >2 fold in copy number. However, interestingly, the copy number of transposons increased with domestication, possibly relecting the genomic plasticity that was required for adaptation under cultivation. Extensive changes in gene expression were seen in developing grains. For example, there was an enrichment for certain functions: genes involved in vesicle traficking in the endosperm showed a gradual increase in expression during durum evolution and genes related to germination and germination inhibition increased in expression in the embryo, in the more recent stages of durum evolution. The approach described here enables better understanding of the genetic events that shaped modern wheat and identiies genes that can be used for crop improvement.

Wild emmer wheat, Triticum turgidum subsp. dicoccoides, (2n = 4x = 28; genome BBAA), the progenitor of domesticated wheat, is genetically very close to durum and common wheat. This wild taxon has many characteristics that would be valuable if transferred to domesticated wheat. An appropriate exploitation of the "wild" genome requires its dissection into segments and evaluation of the contribution of each segment separately to the performance of domesticated wheat. This work describes the production of a series of chromosome-arm substitution lines (CASLs) of wild emmer in the background of the Israeli common wheat cultivar Bethlehem (2n = 6x = 42; genome BBAADD). The identity of the "wild" arm in each CASL was confirmed through the use of RFLP and SSR markers that were polymorphic between the two taxa. The product of this work is a series of true-breeding agronomic lines the breeding value of which can be evaluated under field conditions in different geographic regions. The usefulness of CASLs in studying the improvement of qualitative and quantitative traits in wheat is discussed.

The importance of hybridization and polyploidization in wheat speciation has been recognized for close to a century (Sakamura 1918; Kihara 1919, 1924, 1954; Percival 1921; Sax 1927). Following these pioneering works, it quickly became apparent that polyploid wheats are not the sum of their constituent genomes. This is not unexpected because the nascent hybrids/polyploids are equipped with a complex set of regulatory elements and of copy number variation that originate from two or more divergent genomes and that generate novel types of interactions and dosage effects. Moreover, they have to adjust at the cytological level, at the level of gene expression, and at the protein level. They also have to maintain genome stability through the regulation of meiotic pairing and recombination, the orchestration of cell division, and the silencing of transposons. The recent studies described here provide an impressive account with regard to the extent and the rapid time course at which a new genetic variant was established upon hybridization and polyploidization. We describe here the current knowledge on the changes that occurred in the wheat genome upon allopolyploidization, starting from the early evolutionary and cytological studies to the recent genomic analyses.

The wheat group has evolved through allopolyploidization, namely, through hybridization among species from the plant genera Aegilops and Triticum followed by genome doubling. This speciation process has been associated with ecogeographical expansion and with domestication. In the past few decades, we have searched for explanations for this impressive success. Our studies attempted to probe the bases for the wide genetic variation characterizing these species, which accounts for their great adaptability and colonizing ability. Central to our work was the investigation of how allopolyploidization alters genome structure and expression. We found in wheat that allopolyploidy accelerated genome evolution in two ways: (1) it triggered rapid genome alterations through the instantaneous generation of a variety of cardinal genetic and epigenetic changes (which we termed "revolutionary" changes), and (2) it facilitated sporadic genomic changes throughout the species' evolution (i.e., evolutionary changes), which are not attainable at the diploid level. Our major findings in natural and synthetic allopolyploid wheat indicate that these alterations have led to the cytological and genetic diploidization of the allopolyploids. These genetic and epigenetic changes reflect the dynamic structural and functional plasticity of the allopolyploid wheat genome. The significance of this plasticity for the successful establishment of wheat allopolyploids, in nature and under domestication, is discussed.

The evolvement of duplicated gene loci in allopolyploid plants has become the subject of intensive studies. Most duplicated genes remain active in neoallopolyploids contributing either to a favourable effect of an extra gene dosage or to the build-up of positive inter-genomic interactions when genes or regulation factors on homoeologous chromosomes are divergent. However, in a small number of loci (about 10%), genes of only one genome are active, while the homoeoalleles on the other genome(s) are either eliminated or partially or completely suppressed by genetic or epigenetic means. For several traits, the retention of controlling genes is not random, favouring one genome over the other(s). Such genomic asymmetry is manifested in allopolyploid wheat by the control of various morphological and agronomical traits, in the production of rRNA and storage proteins, and in interaction with pathogens. It is suggested that the process of cytological diploidization leading to exclusive intra-genomic meiotic pairing and, consequently, to complete avoidance of inter-genomic recombination, has two contrasting effects. Firstly, it provides a means for the fixation of positive heterotic inter-genomic interactions and also maintains genomic asymmetry resulting from loss or silencing of genes. The possible mechanisms and evolutionary advantages of genomic asymmetry are discussed.

Speciation via interspecific or intergeneric hybridization and polyploidization triggers genomic responses involving genetic and epigenetic alterations. Such modifications may be induced by small RNAs, which affect key cellular processes, including gene expression, chromatin structure, cytosine methylation and transposable element (TE) activity. To date, the role of small RNAs in the context of wide hybridization and polyploidization has received little attention. In this work, we performed high-throughput sequencing of small RNAs of parental, intergeneric hybrid, and allopolyploid plants that mimic the genomic changes occurring during bread wheat speciation. We found that the percentage of small RNAs corresponding to miRNAs increased with ploidy level, while the percentage of siRNAs corresponding to TEs decreased. The abundance of most miRNA species was similar to midparent values in the hybrid, with some deviations, as seen in overrepresentation of miR168, in the allopolyploid. In contrast, the number of siRNAs corresponding to TEs strongly decreased upon allopolyploidization, but not upon hybridization. The reduction in corresponding siRNAs, together with decreased CpG methylation, as shown here for the Veju element, represent hallmarks of TE activation. TE-siRNA downregulation in the allopolyploid may contribute to genome destabilization at the initial stages of speciation. This phenomenon is reminiscent of hybrid dysgenesis in Drosophila.

Allopolyploidization is the hybridization between two species, followed by genome doubling. This chapter presents a study on wheat hybrids, demonstrating that allopolyploidization induces genome evolution by activating instantaneous radical genomic upheavals through the generation of genetic and epigenetic alterations.

Nuclear DNA amount, determined by the flow cytometry method, in diploids, natural and synthetic allopolyploids, and natural and synthetic autopolyploids of the tribe Triticeae (Poaceae) is reviewed here and discussed. In contrast to the very small and nonsignificant variation in nuclear DNA amount that was found at the intraspecific level, the variation at the interspecific level is very large. Evidently changes in genome size are either the cause or the result of speciation. Typical autopolyploids had the expected additive DNA amount of their diploid parents, whereas natural and synthetic cytologically diploidized autopolyploids and natural and synthetic allopolyploids had significantly less DNA than the sum of their parents. Thus, genome downsizing, occurring during or immediately after the formation of these polyploids, provides the physical basis for their cytological diploidization, that is, diploid-like meiotic behavior. Possible mechanisms that are involved in genome downsizing and the biological significance of this phenomenon are discussed.

Two classes of 5S DNA units, namely the short (containing units of 410 bp) and the long (containing units of 500 bp), are recognized in species of the wheat (the genera Aegilops and Triticum) group. While every diploid species of this group contains 2 unit classes, the short and the long, every allopolyploid species contains a smaller number of unit classes than the sum of the unit classes of its parental species. The aim of this study was to determine whether the reduction in these unit classes is due to the process of allopolyploidization, that is, interspecific or intergeneric hybridization followed by chromosome doubling, and whether it occurs during or soon after the formation of the allopolyploids. To study this, the number and types of unit classes were determined in several newly formed allotetraploids, allohexaploids, and an allooctoploid of Aegilops and Triticum. It was found that elimination of unit classes of 5S DNA occurred soon (in the first 3 generations) after the formation of the allopolyploids. This elimination was reproducible, that is, the same unit classes were eliminated in natural and synthetic allopolyploids having the same genomic combinations. No further elimination occurred in the unit classes of the 5S DNA during the life of the allopolyploid. The genetic and evolutionary significance of this elimination as well as the difference in response to allopolyploidization of 5S DNA and rDNA are discussed.

Recent studies in the genera Aegilops and Triticum showed that allopolyploid formation triggers rapid genetic and epigenetic changes that lead to cytological and genetic diploidization. To better understand the consequences of cytological diploidization, chromosome pairing and seed fertility were studied in S₁, S₂, and S₃ generations of 18 newly formed allopolyploids at different ploidy levels. Results showed that bivalent pairing at first meiotic metaphase was enhanced and seed fertility was improved during each successive generation. A positive linear relationship was found between increased bivalent pairing, improved fertility, and elimination of low-copy noncoding DNA sequences. These findings support the conclusion that rapid elimination of low-copy noncoding DNA sequences from one genome of a newly formed allopolyploid, different sequences from different genomes, is an efficient way to quickly augment the divergence between homoeologous chromosomes and thus bring about cytological diploidization. This facilitates the rapid establishment of the raw allopolyploids as successful, competitive species in nature.

Allopolyploidy accelerates genome evolution in wheat in two ways: 1) allopolyploidization triggers rapid genome alterations (revolutionary changes) through the instantaneous generation of a variety of cardinal genetic and epigenetic changes, and 2) the allopolyploid condition facilitates sporadic genomic changes during the life of the species (evolutionary changes) that are not attainable at the diploid level. The revolutionary alterations, occurring during the formation of the allopolyploid and leading to rapid cytological and genetic diploidization, facilitate the successful establishment of the newly formed allopolyploid in nature. On the other hand, the evolutionary changes, occurring during the life of the allopolyploids, increase the intra-specific genetic diversity, and consequently, increased fitness, adaptability and competitiveness. These phenomena, emphasizing the dynamic plasticity of the allopolyploid wheat genome with regards to both structure and function, are described and discussed in this review.

Nuclear DNA amount (1C) was determined by flow cytometry in the autotetraploid cytotype of Hordeum bulbosum, in the cytologically diploidized autotetraploid cytotypes of Elymus elongatus, Hordeum murinum subsp. murinum and Hordeum murinum subsp. leporinum, in Hordeum marinum subsp. gussoneanum, in their progenitor diploid cytotypes, and in a newly synthesized autotetraploid line of E. elongatus. Several lines collected from different regions of the distribution area of every taxon, each represented by a number of plants, were analyzed in each taxon. The intracytotype variation in nuclear DNA amount of every diploid and autotetraploid cytotype was very small, indicating that no significant changes have occurred in DNA amount either after speciation or after autopolyploid formation. The autotetraploid cytotypes of H. bulbosum and the cytologically diploidized H. marinum subsp. gussoneanum had the expected additive amount of their diploid cytotypes. On the other hand, the cytologically diploidized autotetraploid cytotypes of E. elongatus and H. murinum subsp. murinum and H. murinum subsp. leporinum had considerably less nuclear DNA (10%-23%) than the expected additive value. Also, the newly synthesized autotetraploid line of E. elongatus showed similar reduction in DNA as its natural counterpart, indicating that the reduction in genome size occurred in the natural cytotype during autopolyploidization. It is suggested that the diploid-like meiotic behavior of these cytologically dipolidized autotetraploids is caused by the instantaneous elimination of a large number of DNA sequences, different sequences from different homologous pairs, leading to differentiation of the constituent genomes. The eliminated sequences are likely to include those that participate in homologous recognition and initiation of meiotic pairing. A gene system determining exclusive bivalent pairing by utilizing the differentiation between the two groups of homologues has been presumably superimposed on the DNA reduction process.

Durum and bread wheat need transgenic traits such as herbicide and disease resistance due to recent evolution of herbicide resistant grass weeds and an intractable new strain of stem rust. Transgenic wheat varieties have not been commercialized partly due to potential transgene movement to wild/weedy relatives, which occurs naturally to closely related Aegilops and other spp. Recombination does not occur in the F₁ hybrid between wheat and its relatives due to the presence of the Ph1 gene on wheat chromosome arm 5BL, which acts as a chaperone, preventing promiscuous homoeologous pairing to similar, but not homologous chromosomes of the wild/weedy species. Thus recombination must occur during backcrossing after the wheat Ph1 gene has been eliminated. Based on these findings, we speculate that Ph1 could be used to prevent gene introgression into weedy relatives. We propose two methods to prevent such transgene establishment: (1) link the transgene in proximity to the wheat Ph1 gene and (2) insert the transgene in tandem with the lethal barnase on any chromosome arm other than 5BL, and insert barstar, which suppresses barnase on chromosome arm 5BL in proximity to Ph1. The presence of Ph1 in backcross plants containing 5BL will prevent the homoeologous establishment of barnase coupled to the desired transgene in the wild population. 5BL itself will be eliminated during repeated backcrossing to the wild parent, and progeny bearing the desired transgene in tandem with barnase but without the Ph1-barstar complex will die.

Recent molecular studies in the genera Aegilops and Triticum showed that allopolyploidization (interspecific or intergeneric hybridization followed by chromosome doubling) generated rapid elimination of low-copy or high-copy, noncoding and coding DNA sequences. The aims of this work were to determine the amount of nuclear DNA in allopolyploid species of the group and to see to what extent elimination of DNA sequences affected genome size. Nuclear DNA amount was determined by the flow cytometry method in 27 natural allopolyploid species (most of which were represented by several lines and each line by several plants) as well as 14 newly synthesized allopolyploids (each represented by several plants) and their parental plants. Very small intraspecific variation in DNA amount was found between lines of allopolyploid species collected from different habitats or between wild and domesticated forms of allopolyploid wheat. In contrast to the constancy in nuclear DNA amount at the intraspecific level, there are significant differences in genome size between the various allopolyploid species, at both the tetraploid and hexaploid levels. In most allopolyploids nuclear DNA amount was significantly less than the sum of DNA amounts of the parental species. Newly synthesized allopolyploids exhibited a similar decrease in nuclear DNA amount in the first generation, indicating that genome downsizing occurs during and (or) immediately after the formation of the allopolyploids and that there are no further changes in genome size during the life of the allopolyploids. Phylogenetic considerations of the origin of the B genome of allopolyploid wheat, based on nuclear DNA amount, are discussed.

One of the intriguing issues concerning the dynamics of plant genomes is the occurrence of intraspecific variation in nuclear DNA amount. The aim of this work was to assess the ranges of intraspecific, interspecific, and intergeneric variation in nuclear DNA content of diploid species of the tribe Triticeae (Poaceae) and to examine the relation between life form or habitat and genome size. Altogether, 438 plants representing 272 lines that belong to 22 species were analyzed. Nuclear DNA content was estimated by flow cytometry. Very small intraspecific variation in DNA amount was found between lines of Triticeae diploid species collected from different habitats or between different morphs. In contrast to the constancy in nuclear DNA amount at the intraspecific level, there are significant differences in genome size between the various diploid species. Within the genus Aegilops, the 1C DNA amount ranged from 4.84 pg in A. caudata to 7.52 pg in A. sharonensis; among genera, the 1C DNA amount ranged from 4.18 pg in Heteranthelium piliferum to 9.45 pg in Secale montanum. No evidence was found for a smaller genome size in annual, self-pollinating species relative to perennial, cross-pollinating ones. Diploids that grow in the southern part of the group's distribution have larger genomes than those growing in other parts of the distribution. The contrast between the low variation at the intraspecific level and the high variation at the interspecific one suggests that changes in genome size originated in close temporal proximity to the speciation event, i.e., before, during, or immediately after it. The possible effects of sudden changes in genome size on speciation processes are discussed.

The aim of the present study was to assess the genetic variation in several Israeli and Turkish populations of wild emmer wheat, Triticum turgidum ssp. dicoccoides, the progenitor of most domesticated wheat. Single spikes were collected in 2002 from 60 plants that grew in six different habitats in Ammiad, northeastern Israel (8-12 plants from each habitat), and in 1998 from 56 plants that grew in seven different habitats in Diyarbakir, southeastern Turkey (8 plants from each habitat). Seeds were planted in a nursery and DNA was extracted from every plant and analyzed by the fluorescent-based amplified fragment length polymorphism (AFLP) method. Seven primer combinations produced 788 discernible loci of which 48.6% were polymorphic in Israel and 40.5% in Turkey. The genetic diversity estimates P (frequency of polymorphic loci) and He (gene diversity) were higher in Ammiad than in Diyarbakir (means of P = 0.34 and He = 0.13 in Ammiad vs. P = 0.20 and He = 0.08 in Diyarbakir). Ammiad populations contained more unique alleles than Diyarbakir populations. The relative genetic diversity estimates (θ) values were 0.188 in Ammiad and 0.407 in Diyarbakir, suggesting better differentiation of the populations in Turkey. Genetic distance was larger between Israeli and Turkish populations than between populations of each country. The data indicate that the Israeli and Turkish populations are considerably diverged and that the Israeli populations are more polymorphic than the Turkish ones, having a larger within-populations genetic variation than among-populations one. The significance of the results in relation to the differentiation pattern of wild emmer in the Near East is discussed.

Genetic structure of natural populations of wild crop relatives has been the subject of many studies. Yet, most of them focused on the assessment of spatial genetic diversity, while information on long-term variation, affected by yearly changes, has been considered only in few cases. The present study aimed therefore, to estimate the spatio-temporal genetic variation in populations of wild emmer wheat, the progenitor of domesticated wheat, and to assess the contribution of spatial versus temporal factors to the maintenance of genetic variation in a population. Single spikes were collected in the years 1988 and 2002 from plants that grew in the same sampling points, from six different habitats in the Ammiad conservation site, Eastern Galilee, Israel. Seeds were planted in a nursery and DNA was extracted from each plant and analyzed by the AFLP method. Fourteen primer combinations yielded 1,545 bands of which 50.0 and 48.8% were polymorphic in the years 1988 and 2002, respectively. Genetic diversity was much larger within populations than between populations and the temporal genetic diversity was considerably smaller than the spatial one. Nevertheless, population genetic structure may vary to some degree in different years, mainly due to fluctuations in population size because of yearly rainfall variations. This may lead to predominance of different genotypes in different years. Clustering the plants by their genetic distances grouped them according to their habitats, indicating the existence of genotype-environment affinities. The significance of the results in relation to factors affecting the maintenance of polymorphism in natural populations is discussed.

Methods of reducing stable transgene introgression from cultivated transgenic plants to non-cultivated plants are provided. Specifically, the present invention provides methods of producing transgenic plants having an exogenous genomically integrated sequence that is incapable of stably introgressing into related crops or wild or weedy species.

We describe here the initial steps of cultivation of wild emmer in the Levant, i.e., the western part of the Fertile Crescent, as well as genetic changes caused by spontaneous mutations, leading to its domestication and to the development of free-threshing tetraploid wheat, Triticum turgidum. Review of archaeological findings from the Pre-Pottery Neolithic A (PPNA) (10,300-9,500 BP; uncalibrated) indicates that wild emmer was first cultivated in the southern Levant. Domesticated emmer (with a non-brittle spike) appeared several hundred years later in the early PPNB (9,500-9,000 BP), and for a millennium or more was grown in a mixture with wild emmer in many Levantine sites. After the appearance of domesticated emmer, types with naked, free-threshing grains emerged in the late PPNB (9,000-7,500 BP). We support the model in which domestication occurred independently in several sites across the Levant. According to this view, the genes for non-brittleness were transferred to numerous wild emmer genotypes through countless spontaneous hybridizations, followed by human selection. Consequently, domesticated tetraploid wheat evolved as polymorphic populations rather than single genotypes. The relatively wide genetic basis of the young crop has enabled it to tolerate biotic and abiotic stresses and to succeed under cultivation. The archaeological findings of wild emmer cultivation and domestication do not support the idea of development within a small core area, but rather indicate the polycentric origin of agriculture in the Levant.

Introgressive hybridization has played a crucial role in the evolution of many plant species, especially polyploids. The duplicated genetic material and wide geographical distribution facilitate hybridization and introgression among polyploid species having either homologous or homoeologous genomes. Such introgression may lead to the production of recombinant genomes that are more difficult to form at the diploid level. Crop genes that have introgressed into wild relatives can increase the capability of the wild relatives to adapt to agricultural environments and compete with crops or to compete with other wild species. Although the transfer of genes from crops into their conspecific immediate wild progenitors has been reported, little is known about spontaneous gene movement from crops to more distantly related species. We describe recent spontaneous DNA introgression from domesticated polyploid wheat into distantly related, wild tetraploid Aegilops peregrina (syn. Aegilops variabilis) and the stabilization of this sequence in wild populations despite not having homologous chromosomes. Our results show that DNA can spontaneously introgress between homoeologous genomes of species of the tribe Triticeae and, in the case of crop-wild relatives, possibly enrich the wild population. These results also emphasize the need for fail-safe mechanisms in transgenic crops to prevent gene flow where there may be ecological risks.

Recent studies have shown that allopolyploidy accelerates genome evolution in wheat in two ways: (1) allopolyploidization triggers rapid genome changes (revolutionary changes) through the instantaneous generation of a variety of cardinal genetic and epigenetic alterations, and (2) the allopolyploid condition facilitates sporadic genomic changes during the life of the species (evolutionary changes) that are not attainable at the diploid level. The revolutionary changes comprise (1) non-random elimination of coding and non-coding DNA sequences, (2) epigenetic changes such as DNA methylation of coding and non-coding DNA leading, among others, to gene silencing, (3) activation of genes and retroelements which in tuni alters the expression of adjacent genes. These highly reproducible changes occur in the F₁ hybrids or in the first generation(s) of the nascent allopolyploids and were similar to those that occurred twice in nature: first in the formation of allotetraploid wheat (∼ 0.5 million years ago) and second in the formation of hexaploid wheat (∼ 10,000 years ago). Elimination of non-coding sequences from one of the two homoeologous pairs in tetraploids and from two homoeologous pairs in hexaploids, augments the differentiation of homoeologous chromosomes at the polyploid level, thus providing the physical basis for the diploid-like meiotic behavior of allopolyploid wheat. Regulation of gene expression may lead to improved inter-genomic interactions. Gene inactivation brings about rapid diploidization while activation of genes through demethylation or through transcriptional activation of retroelements altering the expression of adjacent genes, leads to novel expression patterns. The evolutionary changes comprise (1) horizontal intergenomic transfer of chromosome segments between the constituent genomes, (2) production of recombinant genomes through hybridization and introgression between different allopolyploid species or, more seldom, between allopolyploids and diploids, and (3) mutations. These phenomena, emphasizing the plasticity of the genome with regards to both structure and function, might improve the adaptability of the newly formed allopolyploids and facilitate their rapid and successful establishment in nature.

The genomic content of the subtelomeric repeated sequences Spelt1 and Spelt52 was studied by dot, Southern, and in situ hybridization in 11 newly synthesized amphiploids of Aegilops and Triticum, and data were compared with the parental plants. Spelt1 had reduced copy numbers in the first generation of three synthetic amphiploids, but two others did not change; Spelt52 was amplified in nine amphiploids and did not change in two. In the second allopolyploid generation, Spelt1 copy number did not change, whereas there was amplification of Spelt52 in some allopolyploids and decreases in others. Neither allopolyploidy level nor the direction of the cross affected the patterns of change in the newly synthesized amphiploids. Changes did not result from intergenomic recombination because similar alterations were noticed in allopolyploids with and without Ph1, a gene that suppresses homoeologous pairing. No differences in Spelt1 and Spelt52 tandem organization were found by Southern hybridization. The significance of these data are discussed in relation to the establishment of newly formed allopolyploids.

In the past few years we have analysed alterations in genome structure and expression that occur in wheat upon allopolyploidization. Our major findings in natural and synthetic allopolyploid wheat are reviewed here. It was found that allopolyploidization brings about rapid genome evolution through the instantaneous generation of a variety of cardinal genetic and epigenetic alterations comprising: (1) non-random elimination of coding and non-coding DNA sequences, (2) epigenetic changes such as DNA methylation of coding and non-coding DNA leading, among others, to gene silencing, and (3) activation of retroelements, which in turn alters the expression of adjacent genes. These changes were reproducible, occurring in the F1 hybrids or in the first generation(s) of a series of nascent allopolyploids corresponding to various interspecific and intergeneric combinations. Moreover, these changes were similar to those that occurred twice in nature: first, at the transition from diploid to tetraploid wheat (∼0.5 Mya) and, second, at the transition from tetraploid to hexaploid wheat (∼9500 years ago). Elimination of non-coding sequences augments the differentiation of homoeologous chromosomes at the polyploid level, thus increasing the physical divergence between homoeologues and contributing to the diploid-like meiotic behaviour of polyploid wheat. Transcriptional and post-transcriptional alterations of gene activity, including transcriptional activation of retroelements, led to novel expression patterns. These phenomena emphasize the plasticity of the genome with regard to both structure and gene expression. This plasticity in turn might improve the adaptability of the newly formed allopolyploids and facilitate their rapid and successful establishment in nature.

One major strategy to increase the level of zinc (Zn) and iron (Fe) in cereal crops, is to exploit the natural genetic variation in seed concentration of these micronutrients. Genotypic variation for Zn and Fe concentration in seeds among cultivated wheat cultivars is relatively narrow and limits the options to breed wheat genotypes with high concentration and bioavailability of Zn and Fe in seed. Alternatively, wild wheat might be an important genetic resource for enhancing micronutrient concentrations in seeds of cultivated wheat. Wild wheat is widespread in diverse environments in Tarkey and other parts of the Fertile Crescent (e.g., Iran, Iraq, Lebanon, Syria, Israel, and Jordan). A large number of accessions of wild wheat and of its wild relatives were collected from the Fertile Crescent and screened for Fe and Zn concentrations as well as other mineral nutrients. Among wild wheat, the collections of wild emmer wheat, Triticum turgidum ssp. dicoccoides (825 accessions) showed impressive variation and the highest concentrations of micronutrients, significantly exceeding those of cultivated wheat. The concentrations of Zn and Fe among the dicoccoides accessions varied from 14 to 190 mg kg⁻¹ DW for Zn and from 15 to 109 mg kg⁻¹ DW for Fe. Also for total amount of Zn and Fe per seed, dicoccoides accessions contained very high amount of Zn (up to 7 μg per seed) and Fe (up to 3.7 μg per seed). Such high genotypic variation could not be found for phosphorus, magnesium, and sulfur. In the case of modern cultivated wheat, seed concentrations of Zn and Fe were lower and less variable when compared to wild wheat accessions. There was a highly significant positive correlation between seed concentrations of Fe and Zn. Screening different series of dicoccoides substitution lines revealed that the chromosome 6A, 611, and 5B of dicoccoides resulted in greater increase in Zn and Fe concentration when compared to their recipient parent and to other chromosome substitution lines. The results indicate that Triticum turgidum L. var. dicoccoides (wild emmer) is an important genetic resource for increasing concentration and content of Zn and Fe in modern cultivated wheat.

A low-copy, non-coding chromosome-specific DNA sequence, isolated from common wheat, was physically mapped to the distal 19% region of the long arm of chromosome 3B (3BL) of common wheat. This sequence, designated WPG118, was then characterized by Southern hybridization, PCR amplification and sequence comparison using a large collection of polyploid wheats and diploid Triticum and Aegilops species. The data show that the sequence exists in all polyploid wheats containing the B genome and absent from those containing the G genome. At the diploid level, it exists only in Ae. searsii, a diploid species of section Sitopsis, and not in other diploids including Ae. speltoides, the closest extant relative to the donor of the B genome of polyploid wheat. This finding may support the hypothesis that the B-genome of polyploid wheat is of a polyphyletic origin, i.e. it is a recombined genome derived from two or more diploid Aegilops species.

Retrotransposons are a principal component of most eukaryotic genomes, representing roughly 40% of the human genome and 50-80% of some grass genomes. They are usually transcriptionally silent but can be activated under certain stresses. Despite their considerable contribution to genome structure, their impact on the expression of adjacent genes is not well understood. The steady-state transcript levels originating from Wis 2-1A retrotransposons are much higher in newly synthesized wheat amphiploids (two or more diverged genomes in the same nucleus). On activation, both Wis 2-1A long terminal repeats drive the readout synthesis of new transcripts from adjacent sequences including the antisense or sense strands of known genes. Here we report that activation of these antisense or sense transcripts is associated with silencing or activation of the corresponding genes, respectively. These data, together with the abundance of retrotransposons in genomes and their ability to be activated by various signals, support the view of transposons as potential controlling elements.Errata: In the version of this article initially published, Figure 2 was inappropriately enhanced. The original data have been restored to the figure. The authors state that all the display items in this publication are previously unpublished work and accurate representations of the experiments described. The error has been corrected in the HTML and PDF versions of the article.

The present invention provides a method for stably maintaining a genic malesterile female parental line of bread and durum wheat for the production of hybrid wheat. It also provides a male-sterile female line homozygous for a recessive malesterility allele and for a dominant pollen-killer allele, and a maintainer line that is readily and stably propagated. The maintainer line isogenic to the female line but has a stable alien engineered chromosomal arm, added to the wheat complement, carrying a dominant male-fertilizy allele that restores fertility to the maintainer line, a rece ssive pollen-killer allele which is sensitive to the dominant polen-killer allele thus causing abortion of pollen grains carrying it and thus preventing the transmission of this chromosome arm to the femal line, and a heterologous selectable marker that facilitates the selection of maintainer plants among the progeny of the selfed maintainer line and thus, maintenance of the maintainer itself.

The present invention provides a method for stably maintaining a genic male-sterile female parental line of bread and durum wheat for the production of hybrid wheat. It also provides a male-sterile female line homozygous for a recessive male sterility allele, and a maintainer line which is readily and stably propagated. The maintainer line is isogenic to the female line but has an alien chromosomal arm, added to the wheat complement, carrying a dominant male-fertility allele that restores fertility to the maintainer line, a heterologous microspore-suicide gene that kills microspores or pollen grains carrying it thereby preventing the transmission of this chromosome arm to the female line, and a heterologous selectable marker that facilitates the selection of maintainer plants among the progeny of the selfed maintainer line and thus, maintenance of the maintainer itself.

A method for improved production of hybrid seed under genetically (genic or cytoplasmic) induced male sterility of the female line. More specifically, differential genetic response to specific herbicides is used to grow a mixture of a male-sterile female line that is also resistant to the herbicide and a fertile male line that is also susceptible to said herbicide in the production field and spray the mixture of male and female plants with herbicide at a late development stage that will damage the male plants only to prevent normal seed development after pollen shedding by the male plants. The production field is harvested by a combine harvester and the undeveloped male seeds are screened away, and only the normally developed hybrid seeds are harvested. In an alternative method, no herbicide is used.

Recent studies have shown that speciation through allopolyploidy, i.e., inter-specific or inter-generic hybridization followed by chromosome doubling, is accompanied by a variety of rapid cardinal genetic and epigenetic changes. This paper reviews our studies on the effect of allopolyploidization on several low-copy, non-coding sequences that exist in all the diploid species of the tribe Triticeae, including the progenitors of polyploid wheat, but in polyploid wheat they occur in only one genome, either in one homologous pair (chromosome-specific sequences) or in several pairs of the same genome (genome-specific sequences). Rapid elimination of these sequences from one genome is a general phenomenon in newly synthesized allopolyploids. Elimination was a nonrandom, reproducible event whose direction was determined by the genomic combination of the amphiploid. It was not affected by the genotype of the parental plants, by their cytoplasm, or by the ploidy level, and it did not result from intergenomic recombination. This elimination augmented the differentiation of homeologous chromosomes (partially homologous chromosomes of the different genomes) at the polyploid level, thus providing the physical basis for the diploid-like meiotic behavior characterizing polyploid wheat. This pattern of pairing prevents intergenomic recombination, and consequently, ensures full fertility, disomic inheritance, and permanent heterosis between alleles of different genomes (homeoalleles). Accordingly, rapid elimination of these sequences improves the fitness of newly formed allopolyploids, facilitating their rapid establishment in nature as new successful species.

We analyzed the events that affect gene structure and expression in the early stages of allopolyploidy in wheat. The transcriptome response was studied by analyzing 3072 transcripts in the first generation of a synthetic allotetraploid (genome S^lS^lA^mA^m), which resembles tetraploid wheat (genome BBAA), and in its two diploid progenitors Aegilops sharonensis (S^lS^l) and Triticum monococcum ssp. aegilopoides (A^mA^m). The expression of 60 out of 3072 transcripts was reproducibly altered in the allotetraploid: 48 transcripts disappeared and 12 were activated. Transcript disappearance was caused by gene silencing or by gene loss. Gene silencing affected one or both homeologous loci and was associated in part with cytosine methylation. Gene loss or methylation had occurred already in the F₁ intergeneric hybrid or in the allotetraploid, depending on the locus. The silenced/lost genes included rRNA genes and genes involved in metabolism, disease resistance, and cell cycle regulation. The activated genes with a known function were all retroelements. These findings show that wide hybridization and chromosome doubling affect gene expression via genetic and epigenetic alterations immediately upon allopolyploid formation. These events contribute to the genetic diploidization of newly formed allopolyploids.

During the course of our studies on the effect of allopolyploidy on genome evolution in wheat, we produced a large number of different allopolyploids. Single plants used as parents were bagged and selfed so that the genotype of the synthetic allopolyploids can be traced to specific parental plants. The allopolyploids were produced by colchicine treatment and following treatment, plants were grown in the greenhouse until maturity and all spikes were bagged. Three allopolyploids were derived from spontaneous formation of unreduced gametes on F1 plants that were not treated with colchicine. All in all, twenty-three different allopolyploids were obtained (Table 1) of which 10 are "Natural", i.e., they have a genomic combination that exists in nature, and the remainder 13 are "Non-Natural" having a genomic combination that does not exists in nature (Table 1). Chromosome number was determined in all the newly synthesized allopolyploids and only those having the expected euploid chromosome number were harvested. Small samples of seeds, S2, S3, and S4 of all these 23 allopolyploids and of their parental plants are available upon request from the senior author (Hakan Ozkan)

Wild emmer wheat, Triticum turgidum ssp. dicoccoides, the progenitor of most domesticated wheats, was discovered by Aaron Aaronsohn in 1906 in the Levant. The contribution of this discovery to the study of the cytogenetic structure and evolution of the wheats, to an understanding of the processes and site of their domestication and evolution under cultivation, and to the improvement of domesticated wheat is discussed.

To better understand genetic events that accompany allopolyploid formation, we studied the rate and time of elimination of eight DNA sequences in F1 hybrids and newly formed allopolyploids of Aegilops and Triticum. In total, 35 interspecific and intergeneric F1 hybrids and 22 derived allopolyploids were analyzed and compared with their direct parental plants. The studied sequences exist in all the diploid species of the Triticeae but occur in only one genome, either in one homologous pair (chromosome-specific sequences [CSSs]) or in several pairs of the same genome (genome-specific sequences [GSSs]), in the polyploid wheats. It was found that rapid elimination of CSSs and GSSs is a general phenomenon in newly synthesized allopolyploids. Elimination of GSSs was already initiated in F1 plants and was completed in the second or third allopolyploid generation, whereas elimination of CSSs started in the first allopolyploid generation and was completed in the second or third generation. Sequence elimination started earlier in allopolyploids whose genome constitution was analogous to natural polyploids compared with allopolyploids that do not occur in nature. Elimination is a nonrandom and reproducible event whose direction was determined by the genomic combination of the hybrid or the allopolyploid. It was not affected by the genotype of the parental plants, by their cytoplasm, or by the ploidy level, and it did not result from intergenomic recombination. Allopolyploidy-induced sequence elimination occurred in a sizable fraction of the genome and in sequences that were apparently noncoding. This finding suggests a role in augmenting the differentiation of homoeologous chromosomes at the polyploid level, thereby providing the physical basis for the diploid-like meiotic behavior of newly formed allopolyploids. In our view, this rapid genome adjustment may have contributed to the successful establishment of newly formed allopolyploids as new species.

The Ph1 gene has long been considered the main factor responsible for the diploid-like meiotic behavior of polyploid wheat. This dominant gene, located on the long arm of chromosome 5B (5BL), suppresses pairing of homoeologous chromosomes in polyploid wheat and in their hybrids with related species. Here we report on the discovery of genotypic variation among tetraploid wheats in the control of homoeologous pairing. Compared with the level of homoeologous pairing in hybrids between Aegilops peregrina and the bread wheat cultivar Chinese Spring (CS), significantly higher levels of homoeologous pairing were obtained in hybrids between Ae. peregrina and CS substitution lines in which chromosome 5B of CS was replaced by either 5B of Triticum turgidum ssp. dicoccoides line 09 (TTD09) or 5G of Triticum timopheevii ssp. timopheevii line 01 (TIM01). Similarly, a higher level of homoeologous pairing was found in the hybrid between Ae. peregrina and a substitution line of CS in which chromosome arm 5BL of line TTD140 substituted for 5BL of CS. It appears that the observed effect on the level of pairing is exerted by chromosome arm 5BL of T. turgidum ssp. dicoccoides, most probably by an allele of Ph1. Searching for variation in the control of homoeologous pairing among lines of wild tetraploid wheat, either T. turgidum ssp. dicoccoides or T. timopheevii ssp. armeniacum, showed that hybrids between Ae. peregrina and lines of these two wild wheats exhibited three different levels of homoeologous pairing: low, low intermediate, and high intermediate. The low-intermediate and high-intermediate genotypes may possess weak alleles of Ph1. The three different T. turgidum ssp. dicoccoides pairing genotypes were collected from different geographical regions in Israel, indicating that this trait may have an adaptive value. The availability of allelic variation at the Ph1 locus may facilitate the mapping, tagging, and eventually the isolation of this important gene.

Interspecific or intergeneric hybridization, followed by chromosome doubling, can lead to the formation of new allopolyploid species. Recent studies indicate that allopolyploid formation is associated with genetic and epigenetic changes, although little is known about the type of changes that occur, how rapidly they occur, and the type of sequences involved. To address these matters, we have surveyed F1 hybrids between diploid species from the wheat (Aegilops and Triticum) group and their derived allotetraploids by screening a large number of loci using amplified fragment length polymorphism and DNA gel blot analysis and by assaying the extent of cytosine methylation. We found that sequence elimination is one of the major and immediate responses of the wheat genome to wide hybridization or allopolyploidy, that it affects a large fraction of the genome, and that it is reproducible. In one cross between Ae. sharonensis x Ae. umbellulata, 14% of the loci from Ae. sharonensis were eliminated compared with only 0.5% from Ae. umbellulata, with most changes occurring in the F1 hybrid. In contrast, crosses between Ae. longissima x T. urartu showed that sequence elimination was more frequent after chromosome doubling. Alterations in cytosine methylation occurred in ∼13% of the loci, either in the F1 hybrid or in the allopolyploid. For eight of nine bands that were isolated, the sequences that underwent elimination corresponded to low-copy DNA, whereas alterations in methylation patterns affected both repetitive DNA sequences, such as retrotransposons, and low-copy DNA in approximately equal proportions.

NA

The RbcS multigene family of hexaploid (bread) wheat, Triticum aestivum (genome BBAADD), which encodes the small subunit of Rubisco, comprises at least 22 genes. Based on their 3' non-coding sequences, these genes have been classified into four subfamilies (SFs), of which three (SF-2, SF-3 and SF-4) are located on chromosomes of homoeologous group 2 and one (SF-1) on homoeologous group 5. In the present study we hybridized three RbcS subfamily-specific probes (for SF-1, SF-2 and SF-3) to total DNA digested with four restriction enzymes and analyzed the RFLP patterns of these subfamilies in eight diploid species of Aegilops and Triticum, and in two tetraploid and one hexaploid species of wheat (the diploid species are the putative progenitors of the polyploid wheats). The three subfamilies varied in their level of polymorphism, with SF-2 being the most polymorphic in all species. In the diploids, the order of polymorphism was SF-2 > SF3 > SF-1, and in the polyploids SF-2 > SF-1 > SF-3. The RbcS genes of the conserved SF-1 were previously reported to have the highest expression levels in all the wheat tissues studied, indicating a negative correlation between polymorphism and gene expression. Among the diploids, the species with the D and the S genomes were the most polymorphic and the A-genome species were the least polymorphic. The polyploids were less polymorphic than the diploids. Within the polyploids, the A genome was somewhat more polymorphic than the B genome, while the D genome was the most conserved. Among the diploid species with the A genome, the RFLP pattern of T. urartu was closer to that of the A genome of the common wheat cultivar Chinese Spring (CS) than to that of T. monococcum. The pattern in Ae. tauschii was similar to that of the D genome of CS. Only partial resemblance was found between the RFLP patterns of the species with the S genome and the B genome of CS.

An Israeli accession (TTD140) of wild emmer, Triticum turgidum var. dicoccoides, was found resistant to several races of powdery mildew. Inoculation of the chromosome-arm substitution lines (CASLs) of TTD140, in the background of the Israeli common wheat cultivar 'Bethlehem' (BL), with five isolates of powdery mildew revealed that only the line carrying the short arm of chromosome 2B of wild emmer (CASL 2BS) exhibited complete resistance to four of the five isolates. To map and tag the powdery mildew resistance gene, 41 recombinant substitution lines, derived from a cross between BL and CASL 2BS, were used to construct a linkage map at the gene region. The map, which encompasses 69.5 cM of the distal region of chromosome arm 2BS, contains six RFLP markers, a morphological marker (glaucousness inhibitor, Wl¹), and the powdery mildew resistance gene. Segregation ratios for resistance in F₂ of BL x CASL 2BS and in the recombinant lines, combined with the susceptability of F₁ progeny to all tested isolates, indicate that resistance is controlled by a single recessive allele. This allele co-segregated with a polymorphic locus detected by the DNA marker Xwg516, 49.4 cM from the terminal marker Xcdo456. The new powdery mildew resistance gene was designated Pm26.

RFLP diversity in the nuclear genome was estimated within and among Israeli populations of wild emmer wheat (Triticum turgidum var. dicoccoides) from a long-term study site at Ammiad (NE Israel), and from several other geographical locations. Using 55 enzyme-probe combinations, high levels of genetic diversity were revealed in wild emmer in general and within the Ammiad site. In spite of high diversity, observed heterozygosity was low and populations consisted of a patchwork of alternate multilocus homozygotes, consistent with the reproductive biology of a predominant self-fertilizing species. Retention of genetic diversity in wild emmer may be promoted by large population sizes, microhabitat diversity, and occasional gene flow through both pollen and seed. Population genetic structure in wild emmer appears to have been influenced by historical founder events as well as selective factors. Multivariate analyses indicated that individuals tend to cluster together according to their population of origin, and that there is little geographical differentiation among populations. Sampling of 12 domesticated land-races and both primitive and modern cultivars of T. turgidum revealed high levels of diversity and a large number of alleles that were not detected in the wild emmer populations. This may reflect a long-term domestication process in which wild, semi-domesticated, and domesticated types grew sympatrically, continuing introgression from wild populations, and perhaps also gene flow from trans-specific sources.

The RbcS multigene family in hexaploid wheat, Triticum aestivum, is composed of at least 22 genes that were classified into four subfamilies (SFs). In this study, the relative expression of three SFs was analysed, (1) in various plant organs at different developmental stages of one hexaploid line and two tetraploid lines-a wild and a cultivated one-and (2) in fully expanded flag leaves of 16 diploid, 54 tetraploid, and 40 hexaploid lines. The expression of RbcS genes was found to be tissue-specific (highest expression in leaves and stems) and negatively correlated with organs' age. In all cases at all ploidy levels, in all plant organs and developmental stages, and in most light treatments SF-I genes accounted for at least 80% of the RbcS mRNA. Similar to other plant species, wheat RbcS genes were found to be light regulated, with the expression of SF-1 genes being less affected by light than the other two SFs. A negative correlation between the expression level and RFLP was found. Accordingly, the SF with the highest level of expression (SF-1) was the most conserved, suggesting a higher selection pressure for the more active RbcS genes.

The analysis of the pattern of isochromosome pairing allows one to distinguish factors affecting presynaptic alignment of homologous chromosomes from those affecting synapsis and crossing-over. Because the two homologous arms in an isochromosome are invariably associated by a common centromere, the suppression of pairing between these arms (intrachromosome pairing) would indicate that synaptic or postsynaptic events were impaired. In contrast, the suppression of pairing between an isochromosome and its homologous chromosome (interchromosome pairing), without affecting intrachromosome pairing, would suggest that homologous presynaptic alignment was impaired. We used such an isochromosome system to determine which of the processes associated with chromosome pairing was affected by the Ph1 gene of common wheat - the main gene that restricts pairing to homologues. Ph1 reduced the frequency of interchromosome pairing without affecting intrachromosome pairing. In contrast, intrachromosome pairing was strongly reduced in the absence of the synaptic gene Syn-B1. Premeiotic colchicine treatment, which drastically decreased pairing of conventional chromosomes, reduced interchromosome but not intrachromosome pairing. The results support the hypothesis that premeiotic alignment is a necessary stage for the regularity of meiotic pairing and that Ph1 relaxes this alignment. We suggest that Ph1 acts on premeiotic alignment of homologues and homeologues as a means of ensuring diploid-like meiotic behavior in polyploid wheat.

The present invention provides a method for the production of hybrid wheat based on the ability to stably maintaining a genic male-sterile female parental line of common and durum wheat It also provides a male-sterile female line homozygous for a recessive male-sterility allele and for a dominant pollen-killing allele, and a maintainer line which is readily and stably propagated. The maintainer line is isogenic to the female line but has an alien engineered chromosome carrying a dominant male-fertility allele that restores fertility to the maintainer line, a recessive pollen-killing allele that is susceptible to the killing effect of the native pollen killer thus preventing transmission of this chromosome to the female line, and one or more selectable markers that facilitate the maintenance of the maintainer itself. The invention also provides procedures for converting any desired cultivar into male-sterile female line and into a maintainer line.

The cytologically diploid-like meiotic behavior of hexaploid wheat (i.e., exclusive bivalent pairing of homologues) is largely controlled by the pairing homoeologous gene Ph1. This gene suppresses pairing between homoeologous (partially homologous) chromosomes of the three closely related genomes that compose the hexaploid wheat complement. It has been previously proposed that Ph1 regulates meiotic pairing by determining the pattern of premeiotic arrangement of homologous and homoeologous chromosomes. We therefore assume that Ph1 action may be targeted at the interaction of centromeres with spindle microtubules - an interaction that is critical for movement of chromosomes to their specific interphase positions. Using monosomic lines of common wheat, we studied the effect of this gene on types and rates of centromere division of univalents at meiosis. In the presence of the normal two doses of Ph1, the frequency of transverse breakage (misdivision) of the centromere of univalent chromosomes was high in both first and second meiotic divisions; whereas with zero dose of the gene, this frequency was drastically reduced. The results suggest that Ph1 is a trans- acting gene affecting centromere-microtubules interaction. The findings are discussed in the context of the effect of Phi on interphase chromosome arrangement.

NA

Recently, we reported on the characterization of the calmodulin (CaM) gene family in wheat [44]. We classified wheat CaM genes into four subfamilies (SFs) designated SF-1 to SF-4, each representing a series of homoeoallelic loci on the homoeologous chromosomes of the three genomes of common wheat. Here we studied the expression of these wheat CaM genes in the course of wheat development. Northern blot analysis using SF-specific probes revealed differences in SF expression levels in different organs and stages of development. Subsequently, cell-specific expression of CaM SFs was investigated by in situ RNA hybridization. In developing seeds, all CaM SFs showed highest expression in the embryo and less in the aleurone and in the starchy endosperm. In primary roots, all four CaM SFs were expressed in the root cap, meristematic regions and in differentiating cells. During development of the roots, expression gradually decreased. The wheat glutenin gene, which was used as a control throughout our experiments, was found to be expressed in the starchy endosperm but not in the aleurone, embryos or vegetative tissues. In stems, at advanced stages of growth, differences in cell-specific expression of CaM SFs were found. For example, SF-2 was highly expressed in differentiating phloem fibers. Thus, CaM genes in common wheat exhibit a developmentally regulated organ-, tissue-, cell- and SF-specific expression patterns.

We recently reported that allopolyploid formation in wheat was accompanied by rapid nonrandom elimination of specific low-copy, noncoding DNA sequences. These sequences occur in all diploid progenitors but are chromosome- or genome-specific at the polyploid level. To further investigate this phenomenon, we studied nine of these sequences, six chromosome-specific sequences and three genome-specific sequences, all isolated from common wheat. The various sequences were hybridized to DNA derived from nine newly synthesized amphiploids at different ploidy levels and to DNA from their parental lines. Although sequences homologous to the probes occur in all parental lines, a nonrandom loss of hybridization fragments was found at a high frequency in all amphiploids studied. In addition, a 'loss/gain' of a hybridization fragment(s) was noticed in some of the amphiploids at lower frequency. Neither the type nor the frequency of changes was affected by intergenomic recombination or DNA methylation. It is suggested that rapid genomic changes culminated in a 'programmed' pattern of elimination and (or) modification of specific low-copy DNA sequences following allopolyploidization. These events augmented the differentiation of homoeologous chromosomes, thus providing the physical basis for the diploid-like cytological behavior of polyploid wheat.

We recently reported that formation of allopolyploid wheat was accompanied by rapid nonrandom changes in low-copy noncoding DNA sequences. In this report we show that following allopolyploidization, changes also occurred in coding sequences. Genomic DNA of nine different newly synthesized amphiploids of different ploidy levels and their parental lines was digested with five restriction enzymes and probed with 43 coding sequences. The sequences, 19 genomic and 24 cDNA sequences, are group (homoeologous) specific and represent the proximal and distal regions of the short and long arms of the seven homoeologous groups of the Triticeae. We revealed three types of changes: disappearance of a parental hybridization fragment(s), appearance of a novel fragment(s), and simultaneous disappearance of a parental fragment(s) and appearance of a novel fragment(s). No elimination of sequences took place, since in every sequence studied the parental hybridization fragments were present in at least one of the enzyme digests. Variations in pattern among individual plants of the same amphiploid, as well as between several synthetic and natural amphiploids, indicated that at least some of the genomic changes occurred at random. Intergenomic recombination was not the cause of the observed changes. Evidence was obtained, however, that changes were also brought about by DNA methylation. Methylation may cause inactivation of genes or modify their expression levels in some of the newly synthesized amphiploid plants, leading to genetic diploidization and gene-dosage compensation and thus increasing variation among individuals.

To study genome evolution in allopolyploid plants, we analyzed polyploid wheats and their diploid progenitors for the occurrence of 16 low-copy chromosome- or genome-specific sequences isolated from hexaploid wheat. Based on their occurrence in the diploid species, we classified the sequences into two groups: group I, found in only one of the three diploid progenitors of hexaploid wheat, and group II, found in all three diploid progenitors. The absence of group II sequences from one genome of tetraploid wheat and from two genomes of hexaploid wheat indicates their specific elimination from these genomes at the polyploid level. Analysis of a newly synthesized amphiploid, having a genomic constitution analogous to that of hexaploid wheat, revealed a pattern of sequence elimination similar to the one found in hexaploid wheat. Apparently, speciation through allopolyploidy is accompanied by a rapid, nonrandom elimination of specific, low-copy, probably noncoding DNA sequences at the early stages of allopolyploidization, resulting in further divergence of homoeologous chromosomes (partially homologous chromosomes of different genomes carrying the same order of gene loci). We suggest that such genomic changes may provide the physical basis for the diploid-like meiotic behavior of polyploid wheat.

The Ph1 (pairing homoeologous) gene is the major factor that determines the diploid-like chromosome behavior of polyploid wheat. This gene, which is located on the long arm of chromosome 5B (5BL), suppresses homoeologous pairing at meiosis while allowing exclusive homologous pairing. In an effort to tag the specific chromosomal region where this gene is located, we have previously microdissected chromosome arm 5BL from bread wheat and produced a plasmid library by random PCR amplification and cloning. In this work we isolated from this library a 5BL-specific probe, WPG90, and mapped it within the interstitial deleted chromosome fragments carrying Phi in common and durum wheat. A PCR assay of Ph1 based on WPG90 was developed that allows an easy identification of homozygous genotypes deficient for this gene.

Many of the crop plants that helped to found Western civilization have their origin in an arc of land that connects the valleys of the Euphrates and Tigris with the Jordan and which has been termed the Fertile Crescent by Breasted (1938). The progenitors or close relatives of these crop plants survive to this day in wild populations in those areas. Increasing urbanization and modern farming practices, herbicides, etc., are a threat to the richness of these gene pools or their very survival.

Keywords: Biochemistry & Molecular Biology; Cell Biology; Genetics & Heredity

Isolation, physical mapping and polymorphism of chromosome-specific DNA sequences in wheat are reported. Following the microdissection of the long arm of chromosome 5B (5BL) of common wheat, its DNA was amplified by degenerate oligonucleotide-primed PCR and directly cloned into plasmid vectors. Characterization of the chromosome arm library showed that ~55% of the inserts are of low-copy nature. Southern analysis using aneuploid lines of common wheat revealed that five of 11 low-copy inserts analyzed map to chromosome arm 5BL; four of these are 5BL-specific. By deletion mapping, the 5BL-specific sequences were located to sub- chromosome arm regions. Based on the hybridization patterns of three 5BL-specific sequences to DNA from a diverse collection of goat-grass (Aegilops) and wheat (Triticum) species, it was concluded that these sequences emerged at different times in the course of evolution of this group of plant species.

Knowledge of the genetic relationships among wheat lines is important for selection of parents both for conventional breeding and hybrid production. The relatedness of 34 Mediterranean common wheat cultivars was assessed using RFLP analysis and parentage analysis. Cluster analysis based on RFLP data revealed seven major groups. Only in two groups the cultivars shared common parents. No relationship was found between the grouping and the origin of the cultivars. Only a slight negative correlation was found between the relationships based on coefficient of parentage and the RFLP-generated genetic distances. Parentage analysis is considered inferior to RFLP analysis for assessment of relatedness due to lack of accurate pedigree information and to selection during the breeding process which bias the actual relationships.

Calmodulin is a ubiquitous transducer of calcium signals in eukaryotes. In diploid plant species, several isoforms of calmodulin have been described. Here, we report on the isolation and characterization of calmodulin cDNAs corresponding to 10 genes from hexaploid (bread) wheat (Triticum aestivum). These genes encode three distinct calmodulin isoforms; one isoform is novel in that it lacks a conserved calcium binding site. Based on their nucleotide sequences, the 10 cDNAs were classified into four subfamilies. Using subfamily-specific DNA probes, calmodulin genes were identified and the chromosomal location of each subfamily was determined by Southern analysis of selected aneuploid lines. The data suggest that hexaploid wheat possesses at least 13 calmodulin-related genes. Subfamilies 1 and 2 were both localized to the short arms of homoeologous-group 3 chromosomes; subfamily 2 is located on all three homoeologous short arms (3AS, 3BS and 3DS), whereas subfamily 1 is located only on 3AS and 3BS but not on 3DS. Further analysis revealed that Aegilops tauschii, the presumed diploid donor of the D-genome of hexaploid wheat, lacks a subfamily-1 calmodulin gene homologue, whereas diploid species related to the progenitors of the A and B genomes do contain such genes. Subfamily 3 was localized to the short arm of homoeologous chromesomes 2A, 2B and 2D, and subfamily 4 was mapped to the proximal regions of 4AS, 4BL and 4DL. These findings suggest that the calmodulin genes within each subfamily in hexaploid wheat represent homoeoallelic loci. Furthermore, they also suggest that calmodulin genes diversified into subfamilies before speciation of Triticum and Aegilops diploid species.

We report here on the successful painting of a specific plant chromosome within its own genome. Isochromosomes for the long arm of chromosome 5 of the wheat B genome (5BL) were microdissected from first meiotic metaphase spreads of a monoisosomic 5BL line of the common wheat Triticum aestivum cv. Chinese Spring. The dissected isochromosomes were amplified by degenerate oligonucleotide-primed PCR in a single tube reaction. The amplified DNA was used as a complex probe mixture for fluorescent in situ hybridization on first meiotic metaphase spreads of lines carrying 5BL as a distinctive marker. Hybridization signals were observed, specifically, along the entire 5BL. In some of the cells, labeling was also detected in two bivalents, presumably those of the 5B 'homoeologues' (partial homologues) found in common wheat (5A and 5D). The probe also revealed discrete domains in tapetal nuclei at interphase, further supporting the probe's high specificity. These data suggest that chromosome- and homoeologous group-specific sequences are more abundant in 5BL than genome-specific sequences. Chromosome-painting probes, such as the one described here for 5BL, can facilitate the study of chromosome evolution in polyploid wheat.

CD4 molecules interact with class n major histocompatibility complex molecules as a critical costimulatory signal in CD4⁺ cell immune activation. CD4 also recognizes a specific region of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp120 forming a binding site for early stages of HTV-1 infection. We designed two software packages, AUTOMAT and CRITIC, which allowed us to identify similarities between regions of HIV-1 proteins and immunoregulatory protein sequences stored in data banks. In this report we have characterized (i) a pentapeptide, SLWDQ, found in both CD4 and HIV-1 gp120, which surprisingly had remained undetected in these two well-studied molecules until now, and (ii) an HLA sequence corresponding to the putative functional site of H2 I-A. We found that a region of gp120 (residues 254-263) known to be similar to a sequence in HLA class II β chain overlaps this functional region. We showed experimentally that these two CD4 and HLA peptide segments inhibit CD4⁺ cell immune activation. There is strong inhibition (50% up to 80%) of immune activation by SLWDQ-containing gp120 segments and a lesser inhibition by the gp120 HLA-homologous segment. In addition, we found that SLWDQ induced in HIV-1-infected individuals a humoral (antibody) and cellular (cytotoxic T lymphocyte) immune reaction. We propose that these HIV-1 gp120 segments, together with the known CD4-binding region, may contribute to the HIV-1-induced immunosuppression by two mechanisms affecting CD4-HLA interaction during T-cell immune activation: autoimmune reaction toward CD4 and direct interference with the CD4-HLA costimulatory signal inducing CD4⁺ cell anergy with, as a consequence, generation of immunosuppression.

We have designed a computer strategy in order to detect systematically peptidic sites with the potential of interfering with the immune regulatory processes. Applying this software to HIV-1 proteins has led us to unravel a few peptidic sites which could either act directly or be the targets of an auto-immune reaction during HIV-1 infection. We previously reported that the SLWDQ pentapeptide identity with a critical site of CD4 could trigger in HIV-1 infected individuals both an humoral and a cellular autoimmune reaction. In this study, we focused on surprising similitudes unravelled by our software Automat, between HIV-1/2 and another immunoregulatory molecule, the Fas protein which is also called the apoptosis-mediating cell-surface antigen.

The Ph1 (pairing homoeologous) gene of common wheat, the main factor responsible for the cytological diploid-like behavior of this polyploid species, has been a subject of intensive cytogenetic studies in the last three decades. This paper reviews the effects of Ph1 on chromosomal pairing and spindle microtubule dynamics. It presents data supporting the hypothesis that this gene exerts its effect at presynaptic stages, affecting the premeiotic alignment of homologous and homoeologous chromosomes, and thereby controls the regularity and pattern of pairing at meiosis.

Three different 3 noncoding sequences of wheat rubisco small subunit (SSU) genes (RbcS) were used as probes to identify the gene members of different RbcS subfamilies in the common wheat cultivar Chinese Spring (CS). All genes of the wheat RbcS multigene family were previously assigned to the long arm of homoeologous group 5 and to the short arm of homoeologous group 2 chromosomes of cv CS. Extracted DNA from various aneuploids of these homoeologous groups was digested with four restriction enzymes and hybridized with three different 3 noncoding sequences of wheat SSU clones. All RbcS genes located on the long arm of homoeologous group 5 chromosomes were found to comprise a single subfamily, while those located on the short arm of group 2 comprised three subfamilies. Each of the ancestral diploid genomes A, B, and D has at least one representative gene in each subfamily, suggesting that the divergence into subfamilies preceded the differentiation into species. This divergence of the RbcS genes, which is presumably accompanied by a similar divergence in the 5 region, may lead to differential expression of various subfamilies in different tissues and in different developmental stages, in response to different environmental conditions. Moreover, members of one subfamily that belong to different genomes may have diverged also in the coding sequence and, consequently, code for distinguishable SSU. It is assumed that such utilization of the RbcS multigene family increases the adaptability and phenotypic plasticity of common wheat over its diploid progenitors.

The inheritance of grain protein percentage and of grain weight were studied by crossing common and durum wheat cultivars with hexaploid and tetraploid breeding lines that excel in grain protein percentage. All high protein lines were descendants of the tetraploid wild emmer Triticum turgidum var. dicoccoides. One hexaploid cultivar was also crossed with a high-protein var. dicoccoides genotype. All crosses were made between low- and high-protein genotypes and were carried out reciprocally for any combination of genotypes; some of them between genotypes of the same ploidy level and some between hexaploid and tetraploid lines. Weight and protein percentage were determined in selfed and crossed grains that developed on the same spike. Mean weight and protein percentage were also determined in F2 grains of all crosses of the same ploidy level, either tetraploid or hexaploid. At any ploidy level, F1 grains resembled the selfed grains of the mother plant both in grain weight and in grain protein percentage, indicating a major maternal effect on both traits. F2 grains had similar grain weight to the heavy-grained parent, and their protein percentage was close to the midparents value. However, a slight indication of cytoplasmic inheritance of grain protein percentage was found in the comparison between most pairs of F2 reciprocals. The interspecific crosses (hexaploid with tetraploid combinations) yielded shrivelled seeds with highly reduced weight but relatively unchanged protein percentage. Weight reduction in the shrivelled hybrid grains (compared with the selfed ones) was more severe when the mother plant was hexaploid rather than tetraploid. The significance of the different tissues in determining grain weight and protein percentage is discussed.

The genes coding for the Rubisco small subunit (SSU) and for the α-subunit of the Rubisco-binding protein were located to chromosome arms of common wheat. HindIII-digested total DNA from the hexaploid cultivar Chinese Spring and from ditelosomic and nullisomic-tetrasomic lines was probed with these two genes, whose chromosomal location was deduced from the disappearance of or from changes in the relative intensity of the relevant band(s). The Rubisco SSU pattern consisted of 14 bands, containing at least 21 different types of DNA fragments, which were allocated to two homoeologous groups: 15 to the short arm of group 2 chromosomes (4 to 2AS, 7 to 2BS, and 4 to 2DS) and 6 to the long arm of group 5 chromosomes (2 on each of arms 5AL, 5BL, and 5DL). The pattern of the Rubisco-binding protein consisted of three bands, each containing one type of fragment. These fragments were located to be on the short arm of group 2 chromosomes. The restriction fragment length polymorphism (RFLP) patterns of several hexaploid and tetraploid lines were highly conserved, whereas the patterns of several of their diploid progenitors were more variable. The variations found in the polyploid species were mainly confined to the B genome. The patterns of the diploids T. monococcum var. urartu and Ae. squarrosa were similar to those of the A and D genome, respectively, in polyploid wheats. The pattern of T. monococcum var. boeoticum was different from the patterns of the A genome, and the patterns of the diploids Ae. speltoides, Ae. longissima, and Ae. Searsii differed from that of the B genome.

Wild tetraploid wheat, Triticum turgidum var. dicoccoides, is characterized by relatively high grain protein percentage (GPP), up to 30% compared to 9-14% in most cultivated wheats, and low grain yield (GY). The potential of this species to serve as a donor of genes that can increase GPP in cultivated wheats was studied. Analyzing the progeny derived from crosses between var. dicoccoides and various cultivated wheats, four genes were identified that increase GPP: two on the short arm of group-1 chromosomes (1AS and 1BS), one on group-5 and one on group-7 chromosomes. Attempts to transfer the "high-protein" genes into commercial varieties resulted in tetraploid and hexaploid breeding lines that, compared to their parental cultivars, had either higher (1-2 percent) GPP but somewhat lower GY or, alternatively, lower, or even similar GPP but significantly higher GY. In general, GPP and GY were negatively correlated (r=-0.40). Nevertheless, several breeding lines exhibited significantly higher grain protein yield (GPY) per unit area, mainly due to increased GY. To improve the strategy of identifying potentially useful "high protein" or "high yield" genes of wild wheat, to locate them to chromosomes and to evaluate their interactions with the genetic background of cultivated wheats, we are currently producing substitution lines of either whole chromosomes or chromosome arms, as well as introgressed populations in commercial varieties of common wheat. Such substitution lines and introgressed populations will be instrumental in studying the effect of dicoccoides genes in the background of commercial varieties on various quantitative traits.

A number of lessons can be learned from the Ammiad study of wild tetraploid wheat as presented in the foregoing papers.

Variation in the electrophoretic pattern of the high-molecular-weight (HMW) glutenin subunits was studied in the Ammiad population of wild tetraploid wheat, Triticum turgidum var. dicoccoides (genome AABB), during a 5-year period (1983-4 to 1987-8). These storage proteins were analysed following one-dimensional sodium-dodecyl-sulphate polyacrylamide-gel electrophoresis (SDS PAGE), using seeds collected annually from individual plants at 249 defined sampling sites distributed in 11 habitats. Since plants did mt grow at all the sampling points each year, 1108 accessions were analysed altogether. The population was found to be highly polymorphic: the HMW glutenin loci of genome A, Glu-Al-1 and Glu-Al-2, had four and two alleles. respectively, and those of genome B. Glu-Bl-1 and Glu-Bl-2, had five and seven alleles, respectively. The A-genome alleles appeared in 4 combinations, and the B-genome alleles appeared in 12 combinations. There were 18 intergenomic combinations (A and B genotypes), some of which were very rare while others were abundant and distributed along transects in clusters. The spatial distribution of these genotypes was nonrandom, with each of the 11 habitats characterized by different genotype frequencies. Yearly changes in genotypes, mostly occurring in the last 2 years of the study, had little effect on the total frequencies of die various genotypes. A high affinity was found between specific HMW glutenin genotypes and certain habitats. This affinity may have resulted from a random fixation of specific genotypes in different habitats (founder effect) or, alternatively, from natural selection, thus indicating either linkage between HMW glutenin alleles and adaptive genes (hitchhiking effect) or fitness of some of these allele combinations to specific micro-environments.

Among the major crops, cereals constitute about 50% of the annual world protein production used for animal and human consumption (Hanson et al. 1982). Among cereals, wheat production exceeds all other crops, accounting for about 40% of the total protein production. Wheat, therefore, has the major contribution of any single crop to the world protein production (Harlan and Starks 1980). Naturally, the least expensive and most effective means of increasing protein production and upgrading its quality is through the improvement of wheat protein. This chapter deals primarily with the quantitative aspects of high protein wheat.

Grain protein percentage (GPP) was studied in 910 accessions of the wild tetraploid wheat, Triticum turgidum var. dicoccoides, collected from 22 populations representing different ecogeographical conditions in Israel. High values of GPP were found, ranging from 19.7% to 28.0% for population means, and from 14.1% to 35.1% for single accessions. Marginal populations had usually lower GPP and smaller intra-population variation than central ones. Repeated sampling of some central populations for four consecutive years revealed relatively large intra-population fluctuations. A high and significant genetic component of variation was found within and between populations by a nested analysis of variance in two nurseries. However, the regression coefficients of parents vs. offsprings were relatively low, indicating a smaller genetic component of variation which may be accounted for by a significant genotype × environment interaction. No correlation was found between GPP and ecological factors, except for soil type: accessions growing on terra-rossa had higher GPP than those growing on basaltic soil. Accessions with black glumes, or with glabrous auricles, or with large grains exhibited high GPP values. A strategy for collecting accessions with high GPP is presented, and the potential use of high GPP genotypes in breeding programs is discussed.

The genetic control of grain protein percentage (GPP) in the wild tetraploid wheat, Triticum turgidum var. dicoccoides, was determined by crossing four accessions of this taxonomic variety with durum cultivar Inbar, and analyzing the parents, F₁ and F₂ populations. Reciprocal crosses indicated no cytoplasmic effect on GPP. The F₂ variation was continuous in all crosses, showing no transgressive segregation. However, crosses between different accessions of var. dicoccoides showed transgressive segregation indicating the presence of different genes for high GPP in these accessions. Grain protein percentage was mostly codominant with high GPP, showing either no dominance, or a weak dominance. Heritability coefficients (broad sense) ranged from 0.30 to 0.53. Correlation coefficients between GPP and yield components were usually significantly negative, with the exception of the number of spikelets per spike, and in some crosses, grain weight. The number and chromosomal location of genes coding for high GPP were determined by the association between GPP and 27 markers (23 morphological and 4 biochemical markers). For this purpose, the genetic control of these markers, their linkage groups and chromosomal location were studied. At least four loci for high GPP that segregated in the F₂ populations are suggested: one on chromosome arm 1AS, marked by the black glume gene (Bg); one on 1BS, marked by the HMW gliadin locus Gli-B1; one on group 5, marked by the genes for beaked glume (Bkg) and toothed palea (Tp); and one on group 7, marked by the kinky neck gene (Kn). The relationship between GPP and several yield components was studied in a similar manner. In general, loci of markers that correlated positively with high GPP were not correlated with a decrease in yield components. Moreover, several loci of var. dicoccoides were associated with an increase in yield components. The utilization of markers for chromosomal location of genes coding for quantitative traits is compared to the technique of aneuploid analysis, commonly used in wheat. The significance of the above findings for breeding is discussed.

The mode of inheritance, linkage groups, and chromosomal location of 23 morphological and 4 biochemical traits were characterized in the wild tetraploid emmer wheat, Triticum turgidum var. dicoccoides. These traits were described and their mode of inheritance was determined by their segregation in four F₂ populations derived from crosses between four var. dicoccoides accessions and a tetraploid durum cultivar. Linkage groups among the genes encoding for these traits were determined or postulated, and their chromosomal location was deduced by linkage to previously located genes. The genetic control of the following traits was characterized and is first reported here: black keel; hairy leaf sheath; hairy auricles; hairy rachilla; hairy kernel brush; obtuse flag leaf; and curved neck/peduncle. The linkage data indicated that developmentally-related genes tended to occur in clusters.

Polymorphism of the high molecular weight glutenin subunits was studied in 456 accessions of the wild wheat Triticum turgidum var. dicoccoides (2n = 4x = 28; genomes AABB), originating from 21 populations in Israel. A total of 50 different SDS PAGE migration patterns were observed, resulting from the combinations of 15 subunit patterns of the A genome and 24 subunit patterns of the B genome. Most migration patterns consisted of five subunits, varying between three and six. The migration patterns of the A genome had zero to three subunitstwo being most common. The apparent molecular weights (MWs) of the slowest migrating subunit (114, 000 to 103, 500) and of the next in rate of migration (106, 000 to 96, 000) were highly correlated (r = 0·97). Also, both subunits were either present (in most accessions) or absent. In 82.3 per cent of the accessions, the third subunit (MW 76, 000 to 71, 500) was absent, while in 16.9 per cent of the accessions all three subunits of the A genome were absent. The migration patterns of the B genome had one to three subunitsthree being most common. The slowest migrating subunit (99, 500 to 93, 000) was present in almost all cases (99·3 per cent). The MWs of the next two subunits (90, 500 to 82, 000 and 86, 000 to 78, 000, respectively) were highly correlated (r = 0·95). Also, either both subunits were present, as in most cases (94·4 per cent), or absent (5·6 per cent). A nomenclature for the genes encoding for the HMW glutenins is proposed based on the following model: The three subunit groups controlled by each genome are encoded by two genes. In genome A, one gene (Glu-A1-1), with 12 alleles, encodes for the two correlated subunit groups 1Ax and 1Ax; the other gene (Glu-A1-2), with 3 alleles, encodes for the fast-migrating subunit group (1Ay). In genome B, one gene (Glu-B1-1), with 8 alleles, encodes for the slow-migrating subunit group (1Bx), and the other gene (Glu-B1-2), with 10 alleles, encodes for the two correlated subunit groups, 1By and 1By. The polymorphism of the HMW glutenin genes found in var. dicoccoides is much higher than that of cultivated wheats as well as of genes coding for enzymes in var. dicoccoides.

Polymorphism of high molecular weight (HMW) glutenin subunits in 466 accessions of the wild tetraploid wheat Triticum turgidum var. dicoccoides in Israel was characterized with regard to the ecogeographical distribution of the HMW glutenin alleles, both between and within 22 populations, and along transects in a single population. While some populations were monomorphic for all the HMW glutenin loci, namely, Glu-A1-1, Glu-A1-2, Glu-B1-1 and Glu-B1-2, others contained up to four alleles per locus. Intrapopulation variability could be predicted by the geographical distribution: marginal populations tended to be more uniform than those at the center of distribution. The various HMW glutenin alleles tended to be clustered, both at a regional level and within a single population along transects of collection. It is suggested that this clustering is due to selection pressures acting both at a regional and at a microenvironmental level. This was confirmed by the significant correlations found between the MW of subunits encoded by Glu-A1-1 and the populations' altitude, average temperature and rainfall. The possible selective values of seed storage proteins are discussed.

Twenty four B genome aneuploid lines (di-telosomics, nullisomic-tetrasomics and tetrasomics) of Triticum aestivum cv Chinese Spring were used in an analysis of the culture ability and regeneration capability of scutellar calli. Several correlations were found between the presence or absence of specific chromosomes and chromosomal arms of the B genome of common wheat and the growth and differentiation capabilities of these calli. The rate of callus growth decreased only when the long arm of chromosome 6B was not present. The absence of chromosomes 3B and 7B did not result in an apparent change in morphogenetic capability, while the absence of other B genome chromosomes was significantly correlated to changes in the frequency of calli that regenerated plants. The presence of the short arm of chromosome 1B was negatively correlated with regeneration, whereas its long arm is probably required to counteract this effect and to maintain the normal ratio of regeneration. The presence of the chromosomal arm 2BS seemed to be essential for differentiation to shoots. In the absence of the short arms of chromosomes 4B and 5B, the rate of regeneration was slightly reduced. In the absence of the long arm of chromosome 6B there was a marked reduction of the ability of scutellar calli to regenerate plants. The use of additional aneuploid lines belonging to homoeologous group 6 revealed that only calli derived from lines having chromosome 6D in their complement regenerated plants similarly to the euploid control. Culture ability and regeneration capability were also analysed with alloplasmic lines of T. aestivum cv Chris. The lines were derived from five species, representing plasma-types of different phylogenetic distances from plasma-type B of T. aestivum. The results showed that when the endogenous cytoplasm (B-type) was exchanged with T. timopheevii cytoplasm (G-type) there was a significant increase in the regeneration of shoots from the scutellar calli.

Two lines of common wheat cv. Chinese Spring carrying mutant alleles for the Ph2 homoeologous pairing-suppressor gene on chromosome arm 3DS promoting homoeologous pairing in wheat interspecific and intergeneric hybrids have been tested for their mitotic sensitivity to colchicine. Both the ph2a mutation, which corresponds to a fairly long deletion of 3DS, and ph2b, which is either an intragenic change or a very small deletion, as well as the ph2a/ph2b heteroallelic combination and the deficiency for the entire 3DS arm (ditelo 3DL), conditioned a significantly lower sensitivity to colchicine than that determined by the wild-type allele Ph2 (euploid and ditelo 3DS). Observation of both metaphase and anaphse root-tip cell populations, treated with various colchicine concentrations, provided similar results. The degree of spindle disruption in 2 × 10^-4 M colchicine, as measured by the percentage of fully affected metaphases (C type), was significantly reduced in ph2 and Ph2^- genotypes and, consequently, a larger proportion of cells could proceed toward anaphase and also had a regular segregation pattern at this stage. The differential sensitivity of ph2 genotypes to colchicine is in the opposite direction to that previously found for ph1 genotypes, lacking the homoeologous pairing suppressor on chromosome arm 5BL. The ph2 mutation, while promoting homoeologous pairing as ph1 does, decreases spindle sensitivity to colchicine with respect to the wild-type (ph2) allele rather than increasing it, as ph1 does. The observed alteration of spindle sensitivity to colchicine that mutants for structurally unrelated but functionally related genes (Ph1 and Ph2) condition is interpreted as a highly probable coincidence in their cellular target. It is assumed therefore that these loci affect the equilibrium between tubulin and microtubules. Through this effect they presumably determine chromosome positioning in somatic and premeiotic stages, leading to different pairing patterns at meiosis.

Forty-one breeding lines of common wheat, derived from crosses between the Israeli cultivars Miriam and Lakhish and high-protein lines of wild tetraploid wheat, Triticum turgidum var. dicoccoides, were tested for various protein and yield parameters in field trials, under typical agronomic conditions. All lines had a higher grain protein percentage (GPP) than the leading Israeli cultivar Deganit, which was grown as a control. Grain yield (GY) ranged in the breeding lines from a low of 2.44 t/ha to as high as that of Deganit (6.95 t/ha). Despite the weak negative correlation between GPP and GY, several lines excelled both in GPP and in GY. The grain protein yield (GPY) of some of these selected breeding lines was higher than that of Deganit; e.g., 1.19 t/ha in the best line vs. 1.02 t/ha in Deganit. The 16.7% increase in GPY in this line reflected a more efficient utilization of nitrogen.

The polyploid species of the weat (Triticum and Aegilops) group constitute a classical example of evolutionary success through allopolyploidy. The evolutionary advantage of these polyploids over their diploid progenitors reflects a successful genètic system based on allopolyploidy, diploid-like cytological behavior and predominant self-pollination. The different genomes of the newly formed allopolyploids, derived recurrently from diverging diploid species, differ by numerous allelic variations of homoeologous loci. The permanent heterozygosity of the different homoeoalleles facilitated enzyme multiplicity and thereby, wider and greater adaptability. Whereas this genetic multiplication has an evolutionary advantage for loci coding for functional proteins, it may be redundant for others, e.g., multigene families such as rRNA genes or storage protein genes. Activity of all loci in such genes might result in over-production and inefficiency. One should expect, therefore, to find in polyploid wheat regulatory processes involved, on one hand, in the preservation of the activity of favorable gene loci and, on the other hand, in the reduction of the number and activity of the redundant ones. The latter include diploidization (inactivation) and gene dosage compensation (reduced gene expression) processes. The presented evidence indicates that diploidization is a non-random process achieved through mutations or intergenomic suppression. In contrast, gene dosage compensation is a non-specific process determined by several post-transcriptional rate-limiting factors. The evolutionary significance of these regulatory processes is discussed.

Several aneuploid lines and one intervarietal substitution line of the hexaploid wheat Triticum aestivum (2n = 6χ = 42; genomes AABBDD) cv. Chinese Spring were used to study the effects of different doses of chromosomes 1B, 1D, or 1A on the amount of the high molecular weight ('HMW') glutenins and gliadins of endosperm. These homeologous chromosomes carry HMW glutenin and gliadin gene clusters on their long and short arms, respectively. Increasing the dosage of chromosome 1B of Chinese Spring in plants having in their 3n endosperm zero or the normal three doses of the homeologue 1D, as well as in plants carrying in their endosperm one dose of 1B of the cultivar Timstein, had a dual effect: on one hand, a nonlinear increase in the amount of each subunit encoded by the chromosome whose dosage was elevated and, on the other hand, a compensating nonspecific decrease in the amount of other HMW glutenin and gliadin subunits encoded either by the homeoalleles on 1D or by the homoalleles on 1B of Timstein, respectively. Deletion of chromosome arm 1BL, which carries only a few HMW glutenin genes, had no significant effect on the amount of HMW glutenins encoded by 1DL and HMW gliadins encoded by 1DS and 1BS. However, deletion of 1BS or 1DS, each carrying many gliadin genes, caused a significant but nonspecific increase in the HMW glutenins and gliadins encoded by the remaining arms of 1B and 1D. The possible mechanism and evolutionary implications of gene-dosage compensation in polyploid wheat are discussed.

Keywords: Medicine, General & Internal

Five field experiments were conducted during 1982 and 1983 to evaluate the yield response of two lines of common wheat and one line of durum wheat to several rhizospheric bacteria. The two lines tested in the 1982 experiment (a common wheat and a durum wheat) showed a considerable yield response to incoulation with Azospirillum brasilense, with an average significant yield incresae of 23%. In the 1983 experiments, variable yield responses, some of which were significant, were obtained by inoculating three wheat lines (two common wheat and one durum wheat) with Azospirillum or with other rhizospheric bacteria, isolated from various Israeli wild relatives of wheat. A certain specificity between the tested wheat lines, the applied bacteria and the particular local conditions was evident. Inverse relationships were found between the maximum percentage yield increase due to inoculation with the most efficient bacteria and the yield level of the non-inoculated plots.

Several high molecular weight endosperm glutenin subunits, coded by genes located on chromosomes 1A, 1B and 1D of common wheat, Triticum aestivum L. em. Thell., were isolated from excised gel segments and subjected to amino acid analysis and peptide mapping; the latter was carried out following a limited digestion with trypsin, chymotrypsin or Staphylococcus aureus - V8 protease. Generally, all high molecular weight glutenins had a similar amino acid composition but several significant differences were observed in some of them. Both analyses revealed that the structural similarity among the various subunits was related to the homology of the genes coding them: subunits coded by homoalleles, i.e., different alleles of the same gene, were most similar; those coded by homoeoalleles, i.e., alleles of homoeologous genes, were less similar; whereas subunits coded either by alleles of different genes of the same gene cluster, or by nonhomoeoalleles of homoeologous clusters, were the least similar. Several small peptides derived from protease digestion of various subunits had a higher than expected staining intensity indicating that small peptide repeats may be interspersed within the glutenin subunits. The evolutionary course of the high molecular weight glutenins is discussed.

The effect of various chromosomes of Aegilops longissima when added to the common wheat cultivar 'Chinese Spring' was evaluated at two levels of nitrogen fertilization for absolute and relative amount of protein in the grain. All the added chromosomes of Ae. longissima increased protein percentage: protein increase by chromosomes D, C and A averaged 3.8% while that by chromosomes F, E, G and B averaged 1.7%. Addition lines F, D and C had a significantly higher protein weight per grain. On the other hand, lines A, E and G had reduced grain protein weight per grain as compared with that of 'Chinese Spring'. Line C carries the HMW glutenin and some of the gliadin subunits of Ae. longissima. The effect of this line, however, and obviously that of the other lines on protein content was through genes controlling the level of storage protein rather than through genes that code directly for these proteins. Nitrogen fertilization affected protein content and the relative amount of the various protein fractions in a similar manner in every addition line. When high levels of nitrogen fertilization were compared to low ones, the relative amount of the HMW glutenins remained constant while that of HMW gliadins increased and that of the LMW subunits decreased. In contrast to the nitrogen effect, increase in protein content by the addition of longissima chromosomes did not change the relative amounts of the various protein fractions.

Distances between marked pairs of bivalents were determined at first meiotic metaphase of common wheat (2 n=6 x=42; genome AABBDD) by tallying the number of bivalents intervening between the marked bivalents. Bivalents of the same genome tended to lie relatively close to one another, whereas those of different genomes were further apart on the metaphase plate. Within each genome the association of telocentric bivalents representing the two different arms of one chromosome was much more intimate than that of genetically nonrelated bivalents. Among chromosomes of different genomes, homoeologous bivalents were relatively closer to one another than nonrelated ones. These results were more pronounced in cells with circularly arranged bivalents on the metaphase plate than in those with a linear arrangement. The data, consistent with previous findings in cold-treated somatic cells, indicate that a similar pattern of chromosomal arrangement exists in somatic and meiotic cells and that chromosomal arrangement observed in arrested metaphases is also maintained on the untreated, functional spindle system.

The spatial relationships between different types of marked pairs of bivalents or trivalents were studied at first meiotic metaphase of tetraploid wheat (2 n=4 x=28; genome AABB) by tallying the number of intervening bivalents between the marked pair. The study was carried out only in pollen mother cells with circularly arranged metaphases. Telocentric bivalents representing the two different arms of one chromosome were found very close to each other. Nonrelated trivalents of the same genome were significantly closer to each other than nonrelated trivalents of different genomes. Homoeologous trivalents tended to lie relatively close to one another. This pattern of chromosomal arrangement agrees well with previous findings in hexaploid wheat, both in mitotic and meiotic cells. It is concluded that in polyploid wheat the chromosomes of each genome lie relatively close to each other and are spatially separated from those of the other genomes.

Total endosperm proteins extracted from both several common wheat cultivars and some intervarietal substitution lines derived from them were fractionated according to their molecular weight in a high resolution one-dimensional gel electrophoresis. The four donor cultivars and the recipient one - 'Chinese Spring', possessed differentially migrating protein bands in the fractions of high molecular weight (HMW) glutenins and gliadins. Several of these bands were identified for the first time in this study. By utilizing intervarietal substitution lines the control of the HMW glutenins and gliadins by chromosomes of homoeologous group 1 was either reaffirmed or, for the new bands, established. Several HMW gliadin subunits showed a considerable variation in their staining intensity in the intervarietal substitution lines indicating that their expression was dependent on the genetic background.

Five lines of tetraploid wheat were tested for their grain protein content at 10 levels of fertilization ranging from 90 to 2610 mg pure nitrogen per plant. The low levels yielded, in all genotypes,the protein percentage normally obtained under agricultural practice or in the natural habitat. The five lines included: two high protein accessions of the wild wheat, 'l'riticu1n i'ltrgidmn var. dicoccoicles, one clitr1t1n cultivar (Inbar), and the F1 and F6 derivatives of a cross between one of the var. clicoccoides accessions and Inbar. Protein percentage of all genotypes was strongly affected by fertilization, although to a different degree; a significant genotype x fertilization interaction was observed. As a result of that interaction the genetic estimate of dominance ("cl") for protein percentage was found to be significantly affected by the fertilization level: at low levels of fertilization the low protein parent (Inhar) was partially dominant, whereas at high levels - the high protein parent (var. clicoccoicles). At the low levels of fertiliz,ttion, the differences between genotypes were more pronounced than at high levels. Hence, the commonly applied agricultural levels are recommended for any genotypic evaluation ofgermplasm for protein percentage. Heterosis was observed in protein weight per grain and grain weight. Protein ·weight per grain was almost unaffected by the level of fertilization and is therefore suggested as a gootl parameter for breeding wheat with high protein content.

Keywords: Oncology

Acetylene reduction activity (ARA) was measured in cores containing roots of various Israeli wild and cultivated wheat lines colonized by Azospirillum. The inoculated plants were grown under greenhouse or field conditions. Although, no measurable ARA was detected during earlier stages of wheat development, 50-600 nmol C₂H₄ g^-1 dry root h^-1 was measured during heading and flowering stages. By using N yield balance and ¹⁵N dilution techniques, it was found that Triticum aestivum cv. Miriam inoculated with Azospirillum accumulated 20% more N (¹⁴N and ¹⁵N) at the booting stage than did the uninoculated control. This difference in N content became less apparent in grains. No significant ¹⁵N dilution could be found and the contribution of atmospheric N₂ to the N content of grains of inoculated plants was negligible. It was concluded that the potential contribution of biological N₂ fixation to spring wheat cultivation in Israel is very low.

The yield response of a common spring wheat cultivar, Triticum aestivum, to inoculation with Azospirillum brasilense was studied at four levels of N fertilization. Plant yield increased due to the inoculation treatment only at medium and high levels of N fertilization, with a maximum yield increase of about 8.0 per cent at the highest level (approximately 1.0 g of pure N per plant). Yield increase was mostly due to an increase in the number of grains per spike, and at the highest level of fertilization, also due to a higher number of spikes per plant. At all N levels, the inoculation caused an increase of 0.5-1.4 per cent in the number of fertile spikelets per main spike. Grain protein percentage was unaffected by the inoculation, though significantly increase due to the fertilization treatments. The occurrence of maximum yield response at the highest N level, the response by early-determined yield components, i.e. spikelet number, and the unaffected grain protein content are in accord with the suggestion that the contribution of Azospirillum brasilense to wheat yield is not through N₂-fixation.

Twenty different wheat genotypes representing a wide genetic variability, were tested for their yield and yield components response to inoculation with Azospirillum brasilense, at two levels of N fertilization. Only two cultivars responded by a significant yield increase of 7.4 and 8.0 per cent - both at the higher N level. The response reflected an increase in the number of grains per plant added as additional spikes. The importance of the host plant genotype for a successful wheat-bacteria association is discussed.

Reciprocal crosses were made between cultivated wheat (Triticum turgidum var. 'durum') and a high-protein line of wild tetraploid wheat (T. turgidum var. 'dicoccoides'). F₁ grains (on maternal spikes) were very similar to the selfed grains on the maternal parent in protein percentage, weight and protein content. These traits were also analyzed in F₃ grains developed on F₂ spikes of segregating populations derived from reciprocal crosses between the same cultivated parent and another high-protein line of var. 'dicoccoides'. No significant differences in the mean values of these traits were found between the reciprocal crosses, indicating no cytoplasmic effect. It has been concluded that these grain characteristics are largely determined by the maternal plant.

The inheritance of the high molecular weight (HMW) glutenins and of several gliadins controlled, respectively, by the long and short arms of chromosome 1B of common wheat was studied. Analysis was carried out on the progeny of two inter-varietal crosses in which the parental lines possessed differentially migrating subunits as revealed by sodium dodecyl sulphate polyacrylamide gel electrophoresis. No recombination event was detected either within the fraction of the HMW glutenins or among most of the gliadin subunits studied indicating that they are controlled by tightly linked gene clusters. One gliadin subunit (B30) showed 25.5% recombination frequency with the rest of the gliadin subunits and 23.5% recombination frequency with the fraction of the HMW glutenin subunits. It has been concluded that this subunit is controlled by a separate locus (Gld-B6), proximal to the major gliadin gene cluster on the short arm of chromosome 1B. Consequently, the recombination percentage between the glutenin loci and most of the gliadin loci was calculated as 49.0 and the distance in centi-Morgans (cM) as 53.6. The estimated distance in cM is very close to the observed recombination percentage. A genetic map of these storage protein genes is presented.

The arrangements of chromosomes with respect to one another comprise two kinds of spatial relationships: (a) within one gametic set of chromosomes, and (b) between homologous sets of chromosomes. In interspecific hybrids or in allopolyploids spatial relationships also exist between homoeologous (partial homologous) or even nonhomologous sets (Avivi and Feldman, 1980).

The T10 sarcoma, induced in a (C57BL/6J × C3HeB/-FeJ)F1 (H-2b × H-2k) mouse, grows locally (L-T10) and generates spontaneous lung metastases (M-T10). L-T10 cells were found to express the H-2b haplotype, whereas M-T10 expressed both the H-2b and H-2k haplotypes. Most L-T10 cloned cells expressed the H-2b haplotype and were not metastatic. The minority expressed both H-2k and H-2b and were metastatic. Serial transplantations of H-2k-negative clones always ended in spontaneous expression of the H-2k haplotype concomitantly with the acquisition of metastatic potency. The expressed H-2k seemed to be associated with the metastatic properties inasmuch as an H-2b-positive-H-2k-negative clone, which had lost the expressed H-2b and was temporarily H-2 negative, remained nonmetastatic until reexpression of the two haplotyes occurred. Serial transfers of H-2k-positive clones resulted in the maintenance of the expressed H-2k haplotype and the retention of metastatic capacity. A shift toward increased metastatic capacity correlated with H-2k expression occurred during serial transfers of every clone tested. Expression of major histocompatibility complex components, rather than their loss, may potentiate the metastatic capacity of tumor cells.

Endosperm protein subunits of 109 primitive and modern lines of hexaploid wheat, Triticum aestivum L. em. Thell., were fractionated by one-dimensional, high resolution, sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis (PAGE). A wide range of both qualitative and quantitative variation was observed in the fractions of the high molecular weight (HMW) glutenin and gliadin subunits of the different lines. The qualitative variation was expressed in the number of subunits per fraction and in their molecular weight, as determined by the differential rate of migration. The quantitative variation was expressed in the differential staining intensity of several subunits. The widest variation was detected in the HMW glutenin and gliadin subunits controlled by chromosome 1B while a much smaller variation was observed in those subunits controlled by chromosome 1A and further smaller variation in the subunits controlled by 1D. Only a small number of subunits in both fractions was found to be controlled by chromosome 1A indicating that diploidization of endosperm protein genes in common wheat has been non-random. The genetic and evolutionary implications of these findings are discussed.

This study reports the production of clones of human killer T cells grown in the presence of TCGF⁴ following sensitization in MLC against (1) allogeneic cells, (2) autologous B^ebv+ lymphocytes, or (3) autologous lymphoma cells. Sensitization against the tumor cells required the addition of macrophages. The expression of the cytotoxic activity of the cloned T lymphocytes required restimulation with the specific stimulator cells. The cytotoxic activity seemed to be MHCrestricted, since (1) cloned allosensitized CTL lysed their corresponding allogeneic targets, but did not lyse autologous B^ebv+ cells or K562 cells; (2) cloned CTL sensitized against autologous B^ebv+ cells lysed their autologous targets but not allogeneic B^ebv+ targets or K562 cells; and (3) cloned CTL sensitized against autologous Burkitt lymphoma cells lysed their corresponding lymphoma targets or autologous B^ebv+ targets but did not lyse allogeneic lymphoma cells or B^ebv+ cells from the same allogeneic donors. The cloned CTL were homogeneous in expressing the OK T8 molecules and in being negative for T4, T10 or M1. At any given time, 2545% of the cloned cells manifested lytic activity. The ultrastructural properties and cell surface OK T markers were different from those of cloned human NK cells. Emphasis is focused on the differences between the structural, functional and culture characteristics of CTL clones produced by direct isolation of MLC responder cells forming conjugates with specific target cells and those of clones from transformed Tcell lines or from T hybridomas.

Total endosperm protein subunits, extracted from the common wheat cultivar Chinese Spring and from some of its aneuploid lines, were fractionated according to their molecular weight (MW) in an improved high resolution one-dimensional sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis (PAGE). The resolution obtained by this method and, in particular, that of the high molecular weight (HMW) glutenin and gliadin subunits approached that of a previous report in which two-dimensional fractionation system based on charge and MW was used. In the cultivar Chinese Spring, 21 discrete protein bands were resolved and the chromosomes controlling many of them were either reconfirmed, or, in some cases, established. The advantages of this high resolution SDS PAGE technique are discussed.

Brojni genski pulovi divljih srodnika kulturnih biljaka sadrže veliki broj ekonomski značajnih gena koji se mogu preneti u gajene biljke i koristiti za njihovo popravljanje. Da bi se ovi genski pulovi efikasno iskoristili, u poslednjih nekoliko decenija, razvijeni su metodi za prenos gena između vrsta i između rodova različitih gajenih biljaka. Divlji srodnici su opisani po grupama, prema njihovoj genetičkoj srodnosti sa gajenim biljkama. Predloženi su i prodiskutovani odgovarajući postupci za transfer gena. Metodi za transfer udaljenog materijala, koji su ukratko razmatrani u ovom radu, uključuju inkorporiranje celih genoma, pojedinačnih hromozoma, hromozomskih krakova, malih hromozomskih segmenata ili pojedinačnih gena, kao i citoplazmatičnih organela i gena. Tehnike koje uključuju selektivni transfer malih udaljenih hromozomskih segmenata koji nose poželjne gene, ocenjuju se kao najbolji pristup za transfer udaljenih gena.

The frequency of interlocking bivalents at first meiotic metaphase of common wheat Triticum aestivum L., which is normally very low, is significantly increased by raising the dosage (from two to three, four and six) of the Ph1 gene, located on the long arm of chromosome 5B (5BL). In several cells more than three bivalents were interlocked in one chain configuration indicating involvement of non-homoeologous bivalents. Plants with reduced dose (one or zero) of Ph1 also exhibited an increased frequency of interlocking but to a lesser extent than those with high gene dosage. However, chains of more than three interlocked bivalents were never observed in these plants, suggesting that with one or zero doses of Ph1 interlocking is restricted to homoeologous bivalents only. Chromosomal arm 5BS affected interlocking in an opposite manner to 5BL; namely, two and four doses of 5BS markedly reduced interlocking frequency. The modification in the frequency of interlocking bivalents by these genetic manipulations represents the first successful attempt to affect interlocking by genetic means. The results are explained on the basis of the hypothesis that this gene system controls somatic and premeiotic association of both homologous and homoeologous chromosomes.

Spatial relationships between chromosomes of different genomes, both homoeologues (genetically related) and non-homoeologues, were studied in root-tip cells of common wheat, Triticum aestivum L. (2n = 6x = 42) by measuring the distance between cytologically marked chromosomes in cold arrested metaphases. No significant differences were found between the means distances of the different homoeologues. Comparisons of distances between homoeologues with those between their corresponding homologues showed clearly that, in every homoeologous group, the homologous chromosomes were always closer to one another than the homoeologues. Such a pattern of homologous association and relative homoeologous separation in premeiotic cells is believed to facilitate the exclusive pairing of homologues which characterizes the meiosis of common wheat. - No significant differences were found for distances of homoeologues compared with distances of non-homoeologues of different genomes. On the other hand, mean distance between chromosomes of different genomes was significantly greater than that between non-homologues of the same genome. This difference, of course, was much greater if distances between homologues were included in the comparison. Hence, the chromosomes of different genomes are relatively separated from one another. This implies that the three wheat genomes are not intermixed but, rather, tend to occupy different areas of the somatic nucleus. The significance of these intergenomic spatial relationships, their genetic control and cellular mechanism is discussed.

Spatial relationships between chromosomes of the same genome, both homologous and non-homologous, were studied in root-tip cells of common wheat, Triticum aestivum (2n = 6x = 42). Mean distance between members of all the 21 homologous pairs (seven in each of the three genomes) and of 45 out of the 63 possible non-homologous combinations of two (21 in each genome) were determined. To minimize disruption of nuclear chromosomal arrangement, the cells were pretreated with cold temperature either in tap water or in a physiological medium (White solution) and distances between cytologically marked chromosomes were measured at metaphase. Comparison of distances for homologues with those for non-homologues indicated clearly that, within each genome, the homologous chromosomes were significantly closer to one another than were the non-homologues. Distances between homologues were similar in all three genomes, as were distances between non-homologues. The data are consistent with the hypothesis that the chromosomes of each genome of common wheat are arranged in the somatic nucleus in a highly specific ordered pattern. In this hypothetical arrangement, homologous chromosomes are closely associated, while the nonhomologues occupy definite positions with respect to one another. The universality of the phenomenon and its cellular mechanism and biological significance are discussed.

Surgical excision of the local intrafootpad tumor of the 3LL lung carcinoma is followed by accelerated growth of its lung metastases. When, however, splenectomy was performed concomitantly with tumor excision, the acceleration of lung metastases was prevented. In cases where excision of the local tumor took place when it reached large sizes, concomitant splenectomy did not prevent the accelerated growth of the lung metastases. If, however, at these stages of tumor growth splenectomy was performed 3 days prior to the excision of the tumor, it did prevent the accelerated growth of metastases. Intrafootpad reinoculation of tumor cells following tumor excision and splenectomy caused further reduction in metastatic growth. The results suggest the existence of two possible distinct mechanisms which control metastatic growth: the local tumor might exert non-immunologically, an inhibitory effect on its lung metastases, and the spleen, possibly via suppressor lymphocytes, may suppress an immune effector activity against the tumor metastases, an activity which is manifested following splenectomy.

A hormonal mutant of common wheat which is characterized, under certain environmental conditions, by a high percentage of stunted plants was inoculated with a mixture of bacteria of the genus Azospirillum. The inoculation resulted in a reduced number of stunted plants and a significant increase in yield in those plants that escaped stunting and developed normally. The possibility that these changes reflect an improvement in the hormonal balance of the inoculated plants rather than the contribution of fixed nitrogen has been suggested.

The response of 42 commercial and primitive lines of durum and common wheat to inoculation with a mixture of several strains of bacteria of the genus Azospirillum, some of which were isolated from roots of Israeli wild wheats, was studied. Several commercial as well as primitive lines exhibited a significantly higher yield in response to bacterial treatment, due either to an increase in the total number of seeds per plant or to an increase in grain weight. There was no reduction in yield in any of the lines tested. The positive response to the bacterial inoculation was obtained under conditions of both low and high nitrogen levels. The genetic differences between the responding and nonresponding lines as well as the possible mechanism of the bacterial effects are discussed.

Keywords: Medicine, Research & Experimental

Mice were injected from day of birth onward with rabbit antimouse IgM antiserum or purified rabbit antimouse IgM antibodies. These mice completely lacked Igpositive cells or serum Ig, as analyzed by specific fluoresceinated antibodies on the fluorescenceactivated cell sorter (FACSII), by polyclonal B cell mitogens and by specific precipitation in agar. These animals were then primed in vivo by antigen emulsified in complete Freund's adjuvant, and, subsequently, their draining lymph nodes were tested for their T cell proliferative responses in vitro, to the relevant antigen and were found to be severely impaired. However, the antigenpresenting capacity of both spleen cells and thioglycollateinduced peritoneal cells was found to be intact.

The immunogenic capacity of thioglycollateinduced peritoneal macrophages of adult splenectomized animals was compared to that of macrophages of shamoperated controls. Macrophages from splenectomized animals were found to be impaired in their function as antigenpresenting cells, both in the education of virgin initiator T lymphocytes and in the stimulation of antigenspecific T memory cells. Macrophages from splenectomized animals were also severely impaired in their phagocytic capacity, as assessed in an opsonindependent bacterial phagocytosis assay. However, they were not impaired in their ability to pynocytose soluble keyhole limpet hemocyanin. These results indicate that the spleen may play a dicisive role in controlling the differentiation of peritoneal macrophages.

Experiments were made to investigate the effect of four anesthetic drugs that are commonly used in surgical practice on the postoperative growth of mouse tumors in syngeneic recipients. These experiments revealed that some of the anesthetics when applied for surgical excision of the local tumor, strongly accelerated postoperative progression of spontaneous lung metastases produced by the 3LL Lewis lung carcinoma and by the B16 melanoma. Some of the drugs caused the appearance of metastases in organs, such as the liver, in which spontaneous metastases are not usually produced by these tumors. A T10 sarcoma clone that does not produce detectable metastases in immune intact mice even following intravenous injection, did produce metastases when injected into animals treated with pentothal sodium.

Chromosomal arrangement in the interphase nucleus has two main aspects: (1) arrangement of chromosomes with respect to nuclear polarity and to other nuclear components, and (2) arrangement of chromosomes with respect to one another. The latter aspect consists of two main types of spatial relationships; (a) relationships between different members of one chromosomal set, (b) relationships between different chromosomal sets. Data concerning various aspects of chromosomal arrangement in the interphase nucleus are presented and discussed and the genetic control as well as subcellular mechanisms which are involvled in nuclear organization, are elucidated. Evidence is presented indicating that, in common wheat, the gene system that determines the specific pattern of chromosomal arrangement in the nucleus is operating via the microtubular elements of the spindle system. The significance of ordered arrangement of chromosomes in the nucleus for the regularity of genetic activity and chromosomal behavior, is pointed out.

In a previous study we showed that allografts of BN fetal bone, unlike allografts of adult bone, are not rejected by allogeneic recipients of the Lewis strain in spite of the existence of major histocompatibility complex (MHC) incompatibility between donors and hosts. In the present study, we analyzed the relationships existing between the host and fetal tissue that determine graft survival. We found that (1) the fetal BN graft, unlike adult grafts, induces in Lewis recipients a vigorous humoral response consisting mainly in the production of IgG antibody that seems to be directed against antigens of la-like specificities. (2) The BN rats are genetically defective in their capacity to respond to determinants and thus are not capable of producing anti-la antibodies; in accordance, Lewis fetal bone grafts are rejected by the BN recipients. (3) Chondrocytes isolated from fetal mouse bones do express la antigenic determinants. We suggest that the survival of an allogeneic fetal graft in an immunologically intact recipient depends on an active and selective immune response directed against the la components associated with the MHC on the embryonic and fetal cells. On the basis of these notions, we propose that the capacity of la determinants expressed on cells of the embryo, to elicit anti-la and IgG alloantibodies in the pregnant mother, determines the capacity of the embryo to escape rejection by the histoincompatible mother. In a previous study (1, 2) we demonstrated that one can successfully transplant orthotopically, in adult immunologically intact rats, allogeneic fetal bones derived from limbs of rat fetuses and thereby correct massive bone loss. Unlike adult-derived bone grafts, fetal bone grafts underwent processes of rapid ossification, bridging extensive bone defects and supporting the development of bone marrow. The striking conclusion from that work was that, unlike other adult allografts, the fetal bone escapes immune destruction in spite of the strong allogeneic barriers. This escape seemed puzzling in view of our observations that fetal bone grafts are not immunologically inert: we found that the tissue surrounding the allogeneic fetal bone graft became heavily infiltrated by lymphoid cells (1, 2). We had evidence that after transplantation the alloantigens of the fetal bone graft are recognized by T lymphocytes of the recipient, leading to the generation of alloantigen-specific helper T cells (1). In addition, we showed that the recipients of fetal bone grafts (unlike the recipients of adult bone) generate a high titer of specific alloantibodies. Thus, the striking resistance of fetal bone grafts to destruction and rejection by the host in spite of strong allogeneic differences contrasts with the generally accepted principles of transplantation genetics and allograft reactions. The phenomenon seemed puzzling in view of observations demonstrating that chondrocytes do express major histocompatibility antigens (36) in a manner similar to other cells of the body. Our aim, therefore, was to investigate the specific mechanisms involved in permitting the escape of a fetal bone allograft from rejection. This problem is of special interest because of its potential relevance to the escape of the allogeneic embryos and fetuses from destruction by the host immune system. It could then clarify aspects of fetomaternal relationships, as well as relations between an organism and its malignant tissues that carry fetal antigens.

Antigen-fed macrophages were able to induce specific sensitization of unprimed syngeneic lymphocytes in vitro. The sensitized lymphocytes caused specific injury to target cells that carried the relevant antigens. In the present study, we investigated the in vivo activity of lymphocytes sensitized by antigen-fed macrophages. Mouse spleen cells were sensitized by macrophages that had been exposed to the radiation leukemia virus (RadLV). The sensitized lymphocytes, which were enriched for T-cells, were injected to syngeneic normal recipients and 4 days later the mice were challenged with RadLV-induced lymphoma cells. By following tumor growth and survival of mice, we have found that the sensitized lymphocytes protected the recipient mice against lymphoma development if injected 4 days before the tumor cells. The protective activity of the sensitized lymphocytes was radioresistant, but they could not protect irradiated hosts. It is suggested that macrophagemediated in vitro sensitization of lymphocytes induces initiator cells that can protect the recipient host by recruitment of a defensive immune response.

Tests were made to determine whether cell surface tumor antigens of metastases differed from the tumor antigens of the cell population of the local tumor growth. C57BL/6 mouse spleen lymphocytes sensitized against monolayers of the local growth of the 3LL Lewis lung carcinoma (L-3LL) in the presence of syngeneic serum generated lymphocytes that were cytotoxic to L-3LL but significantly less cytotoxic to target cells derived from lung metastases (M-3LL). Lymphocytes sensitized against M-3LL were significantly more cytotoxic against M-3LL than against L-3LL cells. Anti-M-3LL cytotoxic lymphocytes but not anti-L-3LL, admixed with either L-3LL cells or M-3LL tumor cells, when injected into syngeneic recipients reduced lung metastasis significantly. Results indicated that cells with high metastatic capacity and distinct antigenic properties exist within the tumor cell population and that immunoselection might be involved in the production of lung metastases.

A new species of Aegilops named Ae. searsii FELDMAN et. KISLEV, from Judean Mountains and Samaria in Israel and Gilead, Ammon and Moav in Jordan, was recently described. The formal publication of this species as Ae. searsii and as Triticum searsii is at present in press (FELDMAN and KISLEV, in press). It belongs to section Sitopsis ZHUK. (Platystachys EIG). Ae. searsii is most closely related to Ae. longissima SCHWEINF. et MUSCH. Both have a 1-rowed ear which generally disarticulates only very close to its base, and its awns are restricted to the uppermost spikelet. Other common features such as very few fertile florets per spikelet and 2-toothed apex of glume of the lateral spikelets, characterized subsection Emarginata EIG, to which they belong together with Ae. sharonensis EIG and Ae. bicornis (FORSK.) JAUB. et Sp.

Unprimed lymphocytes were sensitized in vitro by incubating them with syngeneic macrophages that had been fed with viral or cellular antigens. The sensitized lymphocytes were tested for their cytotoxic activity against virus-infected and noninfected fibroblasts. The antigenic preparations used for priming the macrophages were either tumor cell-free extracts or supernatants from virus productive cells. Cell-free extracts from the productive RadLV-induced lymphoma cells or the nonproductive radiation-induced lymphoma cells were immunogenic when presented to lymphocytes by macrophages. In contrast, cell-free extracts from normal thymocytes were much less immunogenic, suggesting that the presence of viral associated antigens (VAA) can selectively be detected on lymphoma cells by this assay. Fibroblastic cell lines but not primary fibroblasts were also susceptible to the cytotoxic lymphocytes induced by RadLV-fed macrophages. Primary fibroblasts became susceptible to the sensitized lymphocytes either after infection with the corresponding virus, or if not infected, after several passages in vitro, suggesting that neo-antigens cross-reacting with viral antigens appear during sub-culturing of fibroblasts in vitro. This system makes it possible to detect VAA either as immunogens when presented to lymphocytes by macrophages, or as targets for cytotoxic lymphocytes.

Peripheral blood lymphocytes from normal donors expressed spontaneous cytotoxic activity against human diffuse histiocytic lymphoma cell lines. In the unfractionated state, they could not be further sensitized in vitro against these cell lines. By applying cell separation techniques before culture, subpopulations of lymphocytes were obtained which could be sensitized in vitro and manifested cytotoxic activity against human histiocytic lymphoma cells. Three methods of separation were found effective: E rosette enrichment; elimination of Fc receptor positive cells; and removal of nylon wool adherent cells. Under these conditions, cross-reactive cytotoxicity was observed aganst non-neoplastic lymphoblastoid cell lines, but not against normal lymphocytes.

THE thymus¹ is credited with a central role in the generation of clonal diversity of lymphocytes², and it may perform a critical function in the generation and(or) prevention of autoimmune disorders ³. Less attention has been paid to other, non-immunological thymus functions, such as control of haemopoiesis⁴, development of certain leukaemias⁵, and regulatory roles in the organism's hormonal balance⁶.

Mouse fibroblast monolayers can be used as an immunoadsorbent to separate normal, unsensitized rat lymphocytes with receptors for foreign cell surface antigens. We used this approach to investigate if the specificity of thymusdependent cellular immune reactions is based on a functional diversity of the T lymphocytes recognizing the relevant antigens. T lymphocyte diversity was demonstrated by the following findings. Lymphocytes that are adsorbed to fibroblasts of a given phenotype are relatively restricted in their reactivity to cellular antigen. They strongly react against the adsorbing monolayer type, but their reactivity to another, third party fibroblast type is decreased. Lymphocytes that were preabsorbed on one fibroblast type and transferred to fresh monolayer cultures decreased their reactivity to the absorbing fibroblast type, but could fully react against unrelated third party fibroblasts. Pretreatment of the lymphocytes with relevant subcellular antigen prevents their specific adherence to fibroblast monolayers, and, thus, recognition of cellular antigen. Using mosaic fibroblast monolayers composed of fibroblasts of two unrelated phenotypes, we were able to demonstrate two different lymphocyte populations, each characterized by the capacity to recognize either of the adsorbing fibroblast types.

We investigated whether recognition per se of cell surface antigens by lymphocytes is both necessary and sufficient to trigger sensitization of T cells. We assayed sensitization of normal rat lymphocytes against monolayers of mouse fibroblasts by measuring the acquisition of cytotoxicity against target monolayers syngeneic to the sensitizing monolayer, following 5 days of culturing on living or glutaraldehydefixed fibroblasts. Monolayers of either living or glutaraldehydetreated fibroblasts were used as cellular immunoadsorbents to assay recognition. Glutaraldehyde treatment of mouse fibroblasts did not seem to alter the cell surface antigens detectable by alloantibodies. Yet, rat lymphocytes cultured on such monolayers did not undergo sensitization. This was not due to the lack of feeder effect by the glutaraldehydefixed monolayers, since the addition of living rat fibroblasts, which sustained the survival of the lymphocytes, did not result in sensitization. Not even when living fibroblasts were added to syngeneic fixed monolayers did sensitization of the rat lymphocytes against the monolayer cells occur. Yet, addition of living fibroblasts to fixed allogeneic monolayers resulted in sensitization against the living fibroblasts, but not against the fixed cells. To test whether lymphocytes recognize surface antigens of glutaraldehydetreated monolayers, the capacity of the latter to specifically adsorb lymphocytes possessing receptors for the fibroblast antigen was measured. The nonadhering rat lymphocytes, seeded on glutaraldehydetreated C3H fibroblasts, lost their capacity to become sensitized against fresh C3H cells, but retained their capacity to react against BALB/c fibroblasts. Recognition of C3H antigen did take place, yet this was not sufficient to trigger sensitization. Thus, rat lymphocytes seem to recognize the antigens of fixed monolayers, but cannot respond to them.

Spleen explants from mice tolerant to rabbit serum albumin (RSA) failed to react in vitro to dinitrophenyl (DNP)-RSA; antibodies to DNP were, however, produced by such spleens, when stimulated with α-DNP-poly(Lys). To study the function of T and B cells in recognition of carrier determinants, spleen explants from X-irradiated mice, which had been inoculated with combinations of thymus and bone marrow cells from normal and from RSA tolerant donors, were tested for their reactivity in vitro to the DNP-RSA conjugate. A significant response was obtained only by spleens of mice containing bone marrow and thymus from normal donors. Spleens of mice treated with thymus from tolerant and bone marrow from normal or with thymus from normal and bone marrow from tolerant donors did not respond to DNP-RSA. The absence of the response to DNP-RSA by tolerant B cells combined with normal T cells was unexpected. It could not be attributed to binding of the tolerogen to B cells which would have prevented the interaction with T-cells. Neither could the result be attributed to an inhibition of normal cells by RSA-tolerant B-cells. θ-positive cells in the bone marrow are not the cells controlling the recognition of carrier determinants in the B population, since elimination of θ-positive cells did not affect the reactivity of spleens repopulated with B and T cells. Nor are bone marrow macrophages responsible for the lack of reactivity in spleens containing tolerant B cells, since normal macrophages did not restore reactivity. Hence, the production of antibodies to DNP is based on the recognition of carrier determinants not only by T cells, as previously established, but also by B cells. Whether this indicates a B-B in addition to the T-B cell cooperation is an inviting possibility.

Antigen recognition in transplantation reactions induced in vitro by mouse fibroblasts manifests itself in specific binding of T lymphocytes to membrane antigens of fibroblast monolayers. To deplete graft-versus-host (GVH) reactivity we interacted normal mouse lymphocytes with allogeneic fibroblasts; 4-6% of H-2^d lymphocytes adhered to H-2^k or K-2^b monolayers. The nonadhering cells showed a reduced capacity to bind to cells of the original adsorbing H-2 phenotype. However, adherence to fibroblasts of a different H-2 was not impaired. The capacity of the nonadherent cells to evoke GVH reactivity in vivo was measured by assaying (1) splenomegaly of X-rayed animals injected with the nonadherent lymph node cells; (2) cytotoxicity to target cells of the repopulated spleen cells; and (3) enlargement of the draining lymph nodes in a local GVH reaction following injection of nonadherent spleen cells. All of these test systems showed that GVH reactivity was markedly decreased or eliminated in an immunospecific manner by adsorption. Specific adherence demonstrated the diversity of T lymphocytes with regard to cell receptors for alloantigens. This approach may provide a method for the specific elimination of antihost-reactive cells in the transplantation of bone marrow or lymphoid cells.

Keywords: Immunology; Surgery; Transplantation

Keywords: Immunology; Surgery; Transplantation

Keywords: Allergy; Immunology

The distributions of distances between members of homoeologous pairs was studied in common wheat in root-tip cells and meiocytes. In the majority of the pairs the mean distances were found to be significantly shorter than that calculated for two randomly distributed chromosomes. It could be concluded that homoeologues are loosely associated in somatic cells and more markedly so in meiocytes. However, this association is less intimate than that established previously for homologues. The tests were made in material which carries the association suppressor gene in two doses and the present findings support the assumption that in this dosage the suppressor gene has a smaller effect on homologues than on homoeologues.

IT has now been clearly established (FELDMAN, MELLO-SAMPAYO and SEARS 1966) that homologous chromosomes are closely associated in somatic cells of common wheat, Triticum aestimm L. This association becomes more intimate at late anaphase or early telophase and persists during interphase. It relaxes somewhat at metaphase, presumably due to breakdown of the nuclear membrane and the onset of cell division. Although participation of other chromosomal regions in bringing about somatic association is not rded out, FELDMAN, MELLO-SAMPAYO and SEARS (1966) found that the centromere is primarily responsible for positioning the homologous chromosomes near each other. They consider that the association of homologous chromosomes results, in part, from attachment of their centromeres to the same or closely adjacent sites on the nuclear membrane. This accords well with other published information (for review see COMINGS, 1968) that the interphase chromosomes lie at the periphery of the nucleus attached to the nuclear membrane; for the centromere is presumably the normal place for such attachment to occur. Indeed, there are several reports (BEAMS 1948; VANDERLYN 1948; CARLSON 1956; LETTRE and LETTRE 1959; WOOLAM, MILLEN and FORD 1967; and others) indicating that the centromeres are attached to the nuclear membrane. [1st paragraph]

UPADHYA and SWAMINATHN (Indian Jour. Genet. & Pl. Breed., 1965) studied the mechanism regulating pairing in T. zhukovskyi, a natural autoallohexaploid which arose presumably from the cross T. timopheevi x T. monococcum. In a comparative study of meiosis in the 41- and 42-chromosome hybrids between Chinese Spring Mono-5B and T. zhukovskyi, they found that the number of chromosomes entering into as sociations increased when 5B of Chinese Spring was absent. Thus, they concluded that a gene system similar to that found in chromosome 5B of T. aestivum var. vulgare might not be present in T. zhukovskyi, and, therefore, also not in T. timopheevi.

Natural hybridization and gene flow take place between most of the tetraploid species of Aegilops section Pleionathera, as pointed out by Zohary and Feldman (1962). In Israel, variation patterns in mixed populations of A. variabilis, A. biuncialis, and A. ovata and in mixed populations of A. variabilis and A. triuncialis showed that intermediates and many highly introgressed types occur. In Turkey and Greece less detailed field observations confirmed on a wider scale the common occurrence of natural hybridization between five tetraploid members of the section Pleionathera. The tendency of these tetraploid species to form polymorphic mixed populations consisting of several species increases the frequency of the contact between them and thus facilitates further hybridization. Moreover, the tetraploids which comprise one mixed population have one genome in common that serves as a cytological buffer in the interspecific hybrids and makes the gene exchange possible, while the second genomes differ and can recombine. The great adaptability thereby achieved constitutes an evolutionary advantage.