R. von Bothmer

Phylogenetic analysis of the genus Hordeum using repetitive DNA sequences

A set of six cloned barley (Hordeum vulgare) repetitive DNA sequences was used for the analysis of phylogenetic relationships among 31 species (46 taxa) of the genus Hordeum, using molecular hybridization techniques. in situ hybridization experiments showed dispersed organization of the sequences over all chromosomes of H. vulgare and the wild barley species H. bulbosum, H. marinum and H. murinum. Southern blot hybridization revealed different levels of polymorphism among barley species and the RFLP data were used to generate a phylogenetic tree for the genus Hordeum. Our data are in a good agreement with the classification system which suggests the division of the genus into four major groups, containing the genomes I, X, Y, and H. However, our investigation also supports previous molecular studies of barley species where the unique position of H. bulbosum has been pointed out. In our experiments, H. bulbosum generally had hybridization patterns different from those of H. vulgare, although both carry the I genome. Based on our results we present a hypothesis concerning the possible origin and phylogeny of the polyploid barley species H. secalinum, H. depressum and the H. brachyantherum complex.

Characterization of a family of tandemly repeated DNA sequences in Triticeae.

The recombinant plasmid dpTa1 has an insert of relic wheat DNA that represents a family of tandemly organized DNA sequences with a monomeric length of approximately 340 bp. This insert was used to investigate the structural organization of this element in the genomes of 58 species within the tribe Triticeae and in 7 species representing other tribes of the Poaceae. The main characteristic of the genomic organization of dpTa1 is a classical ladder-type pattern which is typical for tandemly organized sequences. The dpTa1 sequence is present in all of the genomes of the Triticeae species examined and in 1 species from a closely related tribe (Bromus inermis, Bromeae). DNA from Hordelymus europaeus (Triticeae) did not hybridize under the standard conditions used in this study. Prolonged exposure was necessary to obtain a weak signal. Our data suggest that the dpTa1 family is quite old in evolutionary terms, probably more ancient than the tribe Triticeae. The dpTa1 sequence is more abundant in the D-genome of wheat than in other genomes in Triticeae. DNA from several species also have bands in addition to the tandem repeats. The dpTa1 sequence contains short direct and inverted subrepeats and is homologous to a tandemly repeated DNA sequence from Hordeum chilense.

Genetic diversity changes and relationships in spring barley (Hordeum vulgare L.) germplasm of Nordic and Baltic areas as shown by SSR markers

To examine changes in the level of and pattern in variability in 197 Nordic and Baltic spring barley cultivars over time we used 21 mapped barley simple sequence repeats (SSRs). A total number of 191 alleles were found from 22 SSR loci. The number of alleles per locus ranged from 2 to 23, with average of 8.63 107 alleles were rare (frequency <0.05) among the cultivars and only one allele was frequently observed (frequency >0.95). The gene diversity between loci in Nordic and Baltic material varied between 0.033 and 0.891. Average gene diversity was 0.623. The SSR data separated two-rowed and six-rowed cultivars. According to analysis of molecular variance (AMOVA) differentiation in two-rowed vs six-rowed accounted for 23.6% of the total variation. Overall no significant decrease of average gene diversity over time could be found. However, differences were observed when spring barleys from northern (north of ∼58°) and southern (south of ∼58°) parts of the Nordic and Baltic area were compared. For the southern ecogeographical region significant decrease of genetic diversity was observed in the middle of the 20th century, whereas no significant changes in the northern part were found. We found larger differentiation between modern and old cultivars in the South compared to the ones in North parts of the region. The magnitude of changes in genetic diversity differed also with the country of origin. Danish cultivars had a significant decrease in diversity in the middle of century, whereas changes in Finland, Norway and Sweden were not significant.

Elimination and duplication of particular Hordeum vulgare chromosomes in aneuploid interspecific Hordeum hybrids

SummarySeeds formed in crosses Hordeum lechleri (6x) x H. vulgare (2x and 4x), H. arizonicum (6x) x H. v. (2x), H. parodii (6x) x H. v. (2x), and H. tetraploidum (4x) x H. v. (2x) produced plants at high or rather high frequencies through embryo rescue. Giemsa C-banding patterns were used to analyze chromosomal constitutions and chromosomal locations on the methaphase plate. Among 100 plants obtained from H. vulgare (2x) crosses, 32 plants were aneuploid with 2n=29 (1), 28 (3), 27 (13), 26 (5), 25 (4), 24 (4), or 22 (2); 50 were euploid (12 analyzed), and 18 were polyhaploid (5 analyzed). Four plants had two sectors differing in chromosome number. Two of four hybrids with H. vulgare (4x) were euploid and two were aneuploid. Parental genomes were concentrically arranged with that of H. vulgare always found closest to the metaphase centre. Many plants showed a certain level of intraplant variation in chromosome numbers. Except for one H. vulgare (4x) hybrids, this variation was restricted to peripherally located non-H. vulgare genomes. This may reflect a less firm attachment of the chromosomes from these genomes to the spindle. Interplant variation in chromosome numbers was due to the permanent elimination or, far less common, duplication of the centrally located H. vulgare chromosomes in all 34 aneuploids, and in a few also to loss/gain of non-H, vulgare chromosomes. This selective elimination of chromosomes of the centrally located genome contrasts conditions found in diploid interspecific hybrids, which eliminate the peripherally located genome. The difference is attributed to changed “genomic ratios’. Derivatives of various H. vulgare lines were differently distributed among euploid hybrids, aneuploids, and polyhaploids. Chromosomal constitutions of hypoploid hybrids revealed a preferential elimination of H. vulgare chromosomes 1, 5, 6, and 7, but did not support the idea that H. vulgare chromosomes should be lost in a specific order. H. vulgare SAT-chromosomes 6 and 7 showed nucleolar dominance. Aneuploidy is ascribed to the same chromosome elimination mechanism that produces haploids in cross-combinations with H. vulgare (2x). The findings have implications for the utilization of interspecific Hordeum hybrids.

Genetic diversity and structure in a natural Elymus caninus population from Denmark based on microsatellite and isozyme analyses

Abstract. Genetic diversity in a natural Elymus caninus population from Denmark was assessed using isozyme and microsatellite markers. A total of 119 individuals from 46 maternal plants were assayed. Microsatellite loci are shown to display higher levels of variation than isozyme loci. The mean number of alleles per locus was 1.04 for isozymes and 1.38 for microsatellites. The percentage of polymorphic loci for isozymes and microsatellites was 4.7% and 23.6% across the maternal plant, respectively. The genetic diversity at population level was 0.1 for isozymes, and 0.63 for microsatellites. The mean genetic diversity at maternal plant level was 0.027 for isozyme loci and 0.117 for microsatellite loci. The average of total allozyme diversity (HT) was 0.22. The average of total microsatellite diversity was 0.56. Isozyme and microsatellite variation showed the same pattern of differentiation between maternal plants. More than 75% total genetic diversity was found among maternal plants. About 25% total genetic diversity was detected within maternal plants. Ten (22.7%) maternal plants produced heterozygous offspring at allozyme loci, and 30 (68.2%) maternal plants gave heterozygous offspring at microsatellite loci. Both types of markers revealed a relatively high genetic diversity in this population.

Genetic variation and differentiation in Nordic populations of Elymus alaskanus (Scrib. ex Merr.) Löve (Poaceae)

Abstract To gain information on the extent and nature of genetic variation in Elymus alaskanus, levels and distribution of genetic variation were assessed within and among 13 populations originating from Iceland, Norway, Sweden and Russia using allozymes. The results showed that four (30.7%) of the 13 loci were polymorphic within the species, while the mean percentage of polymorphic loci within the populations was 1.9%. The mean number of alleles per locus for the species was 1.8 and 1.02 across the populations. Genetic diversity at the species level was low (Hes=0.135), and mean population diversity was notably lower (Hep=0.005). A high degree of genetic differentiation was observed among populations. The salient points emerging from this study are: (1) statistically significant differences were found in allele frequencies among populations for every polymorphic locus (P<0.001), (2) the high mean coefficient of gene differentiation (GST) showed that 95% of the total allozyme variation was attributable to differences among populations, and (3) relatively high genetic distances between the populations were obtained (mean D=0.16). The Norwegian populations had the highest genetic diversity as compared with the other populations. Geographical comparisons revealed three different groups of populations clearly differentiated, i.e. Scandinavia (Norway and Sweden), Iceland and Russia. Cluster and principal coordinates analyses revealed the same genetic patterns of relationships among populations. Generally, this study indicates that E. alaskanus contains low allozymic variation in its populations. The implications of these results for the conservation of the species are discussed.

Distribution of allozymic alleles and genetic diversity in the American Barley Core Collection

Abstract A survey of allozymic alleles and genetic diversity was made for 151 accessions of the American Barley Core Collection. A total of 25 alleles at ten loci were observed. Two loci were monomorphic. The average diversity index for individual loci ranged between 0.026 and 0.649. Most significant differences in allelic frequency and genetic diversity value were found between spring and winter barley. Spring barley showed a greatly higher average diversity than winter barley (t=2.124, P=0.071). The smallest differences in allelic frequencies and diversity values were observed between the two geographical regions, North and South America. Rare alleles were detected only in a few accessions. Seven rare alleles were associated with spring barley. The genetic similarities among the 151 accessions ranged from 0.20 to 1.00, which showed that a high level of genetic variability exists in this set of core accessions. Cluster analysis and principal coordinate analysis did not give clear-cut separation of different types of barley, but most of the winter barley accessions were closely associated.

Genetic diversity among East Asian accessions of the barley core collection as revealed by six isozyme loci

Abstract Studies of allelic variations at six isozyme loci revealed genetic diversity of 380 East Asian accessions of the Barley Core Collection. Genetic variation was found in both cultivars and landraces in different regions. Allelic variations at the Aco-1 and Aco-2 loci were detected for East Asian barley for the first time. Moreover, the Aco-1 locus displayed the highest genetic diversity among the six loci assayed. Indian cultivars showed the highest diversity, followed by Korean and Chinese cultivars. Landraces from Bhutan and Nepal showed the lowest diversity. Cultivars had generally higher diversity than landraces within as well as among regions. The cluster analysis of genetic identity showed that all landraces from different countries can be placed in one group; the cultivars from Japan, India and Korea each form independent groups. Gpi-1 Gu, Pgd-1 Tj, Aco-1 Si, Ndh-2 D and Aco-2 A were rare alleles found in only a few accessions of 6-rowed barley. The Pgd-2 Tn allele was very rare in East Asian accessions.

Giemsa C-banded karyotypes of diploid and tetraploid Hordeum bulbosum (Poaceae).

The similar-looking basic genomes ofHordeum bulbosum (2x and 4x) have five rather similar metacentric, one submetacentric, and one satellited choromosome. C-banding patterns are characterized by one or two centromeric, or juxtacentromeric, small to larger bands in most chromosomes, by bands at the nucleolar organizers, by small or very small telomeric bands, and by the nearly complete lack of intercalary bands. Banding pattern polymorphism is widespread. Banding patterns supported by chromosome morphology enable identification of homologues, and discrimination between non-homologues inH. bulbosum (2x). The C-banded karyotype ofH. bulbosum (4x) supports an autopolyploid origin, but it was possible to identify only homologues of submetacentrics and SAT-chromosomes.

Phylogenetic analysis of Hordeum marinum Huds. based on nucleotide sequences linked to the vrs1 locus

Abstract. More than 100 accessions of Hordeum marinum ssp. marinum, ssp. gussoneanum 2x and ssp. gussoneanum 4x were studied by analyzing the nuclear DNA homologous to the chloroplast translation elongation factor. PCR amplification using the locus-specific primers generated a 932 bp fragment from ssp. marinum, a 915 bp fragment from ssp. gussoneanum 2x, and 915 bp and 931 bp fragments from ssp. gussoneanum 4x. Sequencing of the entire DNAs showed that the 915-bp fragments of diploids and tetraploids are identical, indicating that the diploid probably is the immediate donor of the 915-bp fragment of tetraploids. Although the donor of the 931-bp fragment was not determined, phylogenetic analysis of the data showed that the clade of the 931-bp fragment was included in the H. marinum complex. A MspI site was diagnostic to distinguish ssp. marinum into two clades with disjunct distributions, the Iberian Peninsula (type 2) and other regions (type 1). Nucleotide variation was revealed in the type 2 accessions but all the other clades were highly uniform.