Eitan Millet

Nuclear DNA amount and genome downsizing in natural and synthetic allopolyploids of the genera Aegilops and Triticum

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, non-coding 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.

Genome size and genome evolution in diploid Triticeae species

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 groups 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.

Genome size in natural and synthetic autopolyploids and in a natural segmental allopolyploid of several Triticeae species

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.

Spatio-temporal genetic variation in populations of wild emmer wheat, Triticum turgidum ssp. dicoccoides, as revealed by AFLP analysis

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.

RESTRICTION FRAGMENT LENGTH POLYMORPHISM IN WILD AND CULTIVATED TETRAPLOID WHEAT

ABSTRACT 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 or...

THE CONTRIBUTION OF THE DISCOVERY OF WILD EMMER TO AN UNDERSTANDING OF WHEAT EVOLUTION AND DOMESTICATION AND TO WHEAT IMPROVEMENT

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.

Genetics of resistance to wheat leaf rust, stem rust, and powdery mildew in Aegilops sharonensis

Aegilops sharonensis (Sharon goatgrass) is a wild relative of wheat and a rich source of genetic diversity for disease resistance. The objectives of this study were to determine the genetic basis of leaf rust, stem rust, and powdery mildew resistance in A. sharonensis and also the allelic relationships between genes controlling resistance to each disease. Progeny from crosses between resistant and susceptible accessions were evaluated for their disease reaction at the seedling and/or adult plant stage to determine the number and action of genes conferring resistance. Two different genes conferring resistance to leaf rust races THBJ and BBBB were identified in accessions 1644 and 603. For stem rust, the same single gene was found to confer resistance to race TTTT in accessions 1644 and 2229. Resistance to stem rust race TPMK was conferred by two genes in accessions 1644 and 603. A contingency test revealed no association between genes conferring resistance to leaf rust race THBJ and stem rust race TTTT or between genes conferring resistance to stem rust race TTTT and powdery mildew isolate UM06-01, indicating that the respective resistance genes are not linked. Three accessions (1644, 2229, and 1193) were found to carry a single gene for resistance to powdery mildew. Allelism tests revealed that the resistance gene in accession 1644 is different from the respective single genes present in either 2229 or 1193. The simple inheritance of leaf rust, stem rust, and powdery mildew resistance in A. sharonensis should simplify the transfer of resistance to wheat in wide crosses.

Distribution of Sharon goat grass (Aegilops sharonensis Eig) in Israel

Sharon goat grass (Aegilops sharonensis Eig) is endemic to the Coastal Plain of Israel and south Lebanon where it grows exclusively on sandy soils. Plants of this species were reported in many locations, but most of these reports are old and many of these locations have undergone urban and agricultural development in recent decades. This study is aimed at determining the current area and mode of distribution of this species. In a systematic survey of the Coastal Plain in Israel, we have located 42 disjunct populations in the area between Acre in the north and Gaza in the south. Physical and ecological data of the populations were recorded. The possibility of diminishing genetic diversity of this species due to extinction of populations is discussed.

Discovery and characterization of two new stem rust resistance genes in Aegilops sharonensis

Key messageWe identified two novel wheat stem rust resistance genes, Sr-1644-1Sh and Sr-1644-5Sh in Aegilops sharonensis that are effective against widely virulent African races of the wheat stem rust pathogen.AbstractStem rust is one of the most important diseases of wheat in the world. When single stem rust resistance (Sr) genes are deployed in wheat, they are often rapidly overcome by the pathogen. To this end, we initiated a search for novel sources of resistance in diverse wheat relatives and identified the wild goatgrass species Aegilops sharonesis (Sharon goatgrass) as a rich reservoir of resistance to wheat stem rust. The objectives of this study were to discover and map novel Sr genes in Ae. sharonensis and to explore the possibility of identifying new Sr genes by genome-wide association study (GWAS). We developed two biparental populations between resistant and susceptible accessions of Ae. sharonensis and performed QTL and linkage analysis. In an F6 recombinant inbred line and an F2 population, two genes were identified that mapped to the short arm of chromosome 1Ssh, designated as Sr-1644-1Sh, and the long arm of chromosome 5Ssh, designated as Sr-1644-5Sh. The gene Sr-1644-1Sh confers a high level of resistance to race TTKSK (a member of the Ug99 race group), while the gene Sr-1644-5Sh conditions strong resistance to TRTTF, another widely virulent race found in Yemen. Additionally, GWAS was conducted on 125 diverse Ae. sharonensis accessions for stem rust resistance. The gene Sr-1644-1Sh was detected by GWAS, while Sr-1644-5Sh was not detected, indicating that the effectiveness of GWAS might be affected by marker density, population structure, low allele frequency and other factors.

Exploitation of Aegilops species of section Sitopsis for wheat improvement

Section Sitopsis of the genus Aegilops is comprised of five diploid species sharing the S genome, which is closely related to the B genome of durum and bread wheat. Species of this section are a rich source of economically valuable traits for wheat improvement and, especially, resistance to different wheat diseases. The most exploitable species is Ae. speltoides, which is genetically the closest to the B genome and contains a variety of disease-resistance genes. Many lines of Aegilops speltoides carry a gene that promotes homoeologous pairing between Ae. speltoides and wheat chromosomes. This gene, easing the transfer of genes to wheat, was extensively used in the exploitation of Ae. speltoides genes. Transferring of desired genes from other Sitopsis species is facilitated by the use of the mutant ph1 gene, which allows for pairing between homoeologous chromosomes of these species and those of wheat. Gametocidal genes, interfering with successful gene transfer, are present in Ae. sharonensis, Ae. longissi...