Specific Simple Sequence Repeat Research Articles

The Complete Plastid Genome Sequence of Madagascar Periwinkle Catharanthus roseus (L.) G. Don: Plastid Genome Evolution, Molecular Marker Identification, and Phylogenetic Implications in Asterids.

The Madagascar periwinkle ( Catharanthus roseus in the family Apocynaceae) is an important medicinal plant and is the source of several widely marketed chemotherapeutic drugs. It is also commonly grown for its ornamental values and, due to ease of infection and distinctiveness of symptoms, is often used as the host for studies on phytoplasmas, an important group of uncultivated plant pathogens. To gain insights into the characteristics of apocynaceous plastid genomes (plastomes), we used a reference-assisted approach to assemble the complete plastome of C . roseus , which could be applied to other C . roseus -related studies. The C . roseus plastome is the second completely sequenced plastome in the asterid order Gentianales. We performed comparative analyses with two other representative sequences in the same order, including the complete plastome of Coffea arabica (from the basal Gentianales family Rubiaceae) and the nearly complete plastome of Asclepias syriaca (Apocynaceae). The results demonstrated considerable variations in gene content and plastome organization within Apocynaceae, including the presence/absence of three essential genes (i.e., accD, clpP, and ycf1) and large size changes in non-coding regions (e.g., rps2-rpoC2 and IRb-ndhF). To find plastome markers of potential utility for Catharanthus breeding and phylogenetic analyses, we identified 41 C . roseus -specific simple sequence repeats. Furthermore, five intergenic regions with high divergence between C . roseus and three other euasterids I taxa were identified as candidate markers. To resolve the euasterids I interordinal relationships, 82 plastome genes were used for phylogenetic inference. With the addition of representatives from Apocynaceae and sampling of most other asterid orders, a sister relationship between Gentianales and Solanales is supported.

Registration of Three Germplasm Lines of Cotton Derived from Gossypium barbadense L. Accession GB713 with Resistance to the Reniform Nematode

Three upland cotton (Gossypium hirsutum L.) germplasm lines, M713 Ren1 (Reg. No. GP‐958, PI 665928), M713 Ren2 (Reg. No. GP‐959, PI 665929), and M713 Ren5 (Reg. No. GP‐960, PI 665930) resistant to the reniform nematode, Rotylenchulus reniformis Linford and Oliveria were developed and jointly released by the USDA‐ARS and the Mississippi Agricultural and Forestry Experiment Station in 2012. The day‐neutral, reniform‐resistant germplasm lines originated from the photoperiodic G. barbadense L. accession GB713 and were selected using the tightly linked simple sequence repeat (SSR) markers GH 132, BNL 3279, and BNL 569. Egg production of the reniform nematode was suppressed to approximately 90% below that of the susceptible check, ‘Deltapine 61’ in greenhouse tests. The lines exhibit considerable differences for agronomic and fiber traits such as length, strength and micronaire, and with their high level of resistance they should be valuable to cotton breeding programs. The successful use of marker assisted selection for these specific SSR markers further validates their use in the selection of resistant plants in segregating generations.

Assessment of molecular diversity and evolutionary relationship of kenaf (Hibiscus cannabinus L.), roselle (H. sabdariffa L.) and their wild relatives

Kenaf (Hibiscus cannabinus L.) and roselle (H. sabdariffa L.) are valuable fibre crop species with diverse end use. Phylogenetic relationship of 73 accessions of kenaf, roselle and their wild relatives from 15 countries was assessed using 44 inter-simple sequence repeat (ISSR) and jute (Corchorus olitorius L.) specific simple sequence repeats (SSR) markers. A total of 113 alleles were identified of which 61.95 % were polymorphic. Jute specific SSR markers exhibited high polymorphism and resolving power in kenaf, although ISSR markers exhibited higher resolving power than SSR markers. Number of polymorphic alleles varied from 1 to 5 for ISSR and 1 to 6 for SSR markers. Cultivated species exhibited higher allele polymorphism (57 %) than the wild species (35 %), but the improved cultivars exhibited lower genetic diversity compared to germplasm accessions. Accessions with common genetic lineage and geographical distribution clustered together. Indian kenaf varieties were distinct from cultivars bred in other countries and shared more genetic homology with African accessions. High genetic diversity was observed in the Indian (J = 0.35–0.74) and exotic kenaf germplasm collections (J = 0.38–0.79), suggesting kenaf might have been introduced in India from Africa through Central Asia during early domestication. Genetic similarity-based cluster analysis was in close accordance with taxonomic classification of Hibiscus.

Rapid Microsatellite Identification from Illumina Paired-End Genomic Sequencing in Two Birds and a Snake

Identification of microsatellites, or simple sequence repeats (SSRs), can be a time-consuming and costly investment requiring enrichment, cloning, and sequencing of candidate loci. Recently, however, high throughput sequencing (with or without prior enrichment for specific SSR loci) has been utilized to identify SSR loci. The direct “Seq-to-SSR” approach has an advantage over enrichment-based strategies in that it does not require a priori selection of particular motifs, or prior knowledge of genomic SSR content. It has been more expensive per SSR locus recovered, however, particularly for genomes with few SSR loci, such as bird genomes. The longer but relatively more expensive 454 reads have been preferred over less expensive Illumina reads. Here, we use Illumina paired-end sequence data to identify potentially amplifiable SSR loci (PALs) from a snake (the Burmese python, Python molurus bivittatus), and directly compare these results to those from 454 data. We also compare the python results to results from Illumina sequencing of two bird genomes (Gunnison Sage-grouse, Centrocercus minimus, and Clark's Nutcracker, Nucifraga columbiana), which have considerably fewer SSRs than the python. We show that direct Illumina Seq-to-SSR can identify and characterize thousands of potentially amplifiable SSR loci for as little as $10 per sample – a fraction of the cost of 454 sequencing. Given that Illumina Seq-to-SSR is effective, inexpensive, and reliable even for species such as birds that have few SSR loci, it seems that there are now few situations for which prior hybridization is justifiable.

Computational Analyses of Simple Sequence Repeats on Human Tissue Specific Genes Promoters

Promoter region of gene closely related with tissue specific expression and SSRs (simple sequence repeats) have been shown to have a variety of effects on an organism. This paper used a heuristic method to find SSRs and compared the most frequently SSRs on promoter region of both human tissues specific genes and human housekeeping genes. We used kidney and testis tissue as examples to show the final results. Especially, we found that (AGG)n is kidney specific SSR and (GCG)n is testis specific SSR. We also analyzed the SSRs frequency density distribution on different promoter regions of both tissue specific genes and housekeeping genes, and we found the density of housekeeping genes on core-promoter region is much higher than on other promoter regions.

Persistent C genome chromosome regions identified by SSR analysis in backcross progenies between <i>Brassica juncea</i> and <i>B. napus</i>

Given that feral transgenic canola (Brassica napus) from spilled seeds has been found outside of farmer’s fields and that B. juncea is distributed worldwide, it is possible that introgression to B. juncea from B. napus has occurred. To investigate such introgression, we characterized the persistence of B. napus C genome chromosome (C-chromosome) regions in backcross progenies by B. napus C-chromosome specific simple sequence repeat (SSR) markers. We produced backcross progenies from B. juncea and F1 hybrid of B. juncea × B. napus to evaluate persistence of C-chromosome region, and screened 83 markers from a set of reported C-chromosome specific SSR markers. Eighty-five percent of the SSR markers were deleted in the BC1 obtained from B. juncea × F1 hybrid, and this BC1 exhibited a plant type like that of B. juncea. Most markers were deleted in BC2 and BC3 plants, with only two markers persisting in the BC3. These results indicate a small possibility of persistence of C-chromosome regions in our backcross progenies. Knowledge about the persistence of B. napus C-chromosome regions in backcross progenies may contribute to shed light on gene introgression.

Allelic Diversity Among Short Duration Maize (Zea mays L.) Genotypes Using SSR Markers

In the present investigation, 25 lines of maize were analysed for allelic diversity using 15 carotenoids specific simple sequence repeats (SSRs) markers including gene targeted marker y1SSR. All the markers were found to be polymorphic thus able to differentiate different genotypes. Total of 66 alleles were detected across the 15 polymorphic loci with an average of 4.4 alleles per locus. Polymorphic information content (PIC), a measure of diversity analysis, ranged from 0.63 (umc 2047) to 0.83 (y1SSR). Jaccard’s similarity coefficient was found to vary from 0.16 (between CM-300 and LPP-1) to 0.65 (between Amar and Pragati). The unweighted pair group method with arithmetic mean (UPGMA) dendrogram generated using Jaccard’s similarity coefficients of SSR markers data divided 25 genotypes into two broad groups. Both the groups were further sub-divided into seven clusters. Molecular marker based diversity did not correspond to quantitative estimates of kernel carotenoids.

Additional microsatellite DNA markers for varietal identification and detection of adulteration in Basmati rice

Availability of markers that distinguish commercially important premium Basmati rice varieties from cheaper Basmati or non-Basmati indica rice varieties is of interest to exporters, commercial suppliers and consumers to ensure the varietal purity of Basmati rice supplies. In this study, a fingerprint database of 27 rice varieties including commercially important traditional Basmati (TB), cross-bred (evolved) Basmati and some non-Basmati indica rice varieties has been prepared using 50 microsatellite markers. A total of 271 electromorphs (alleles) were detected at 50 SSR (simple sequence repeat) loci, 92 of which were observed only in 2–10 of the 16 Basmati rice varieties. Notably, 7–10 of the sixteen Basmati rice varieties had the same Basmati specific electromorphs at RM562, RM551 and RM547 loci. Alarge number of unique (22) and null (16) electromorphs were noticed in Basmati rice varieties. A combination of four SSR markers (RM224, RM248, RM547 and RM594) can be used to differentiate all the 27 rice varieties. Specific SSR markers have been identified for the authentication of Premium Basmati rice cultivars such as Taraori Basmati (RM547, RM594 and RM511), Basmati 370 (RM252, RM426 and RM527), Karnal Local (RM248, RM423 and RM488) and Pusa 1121 (RM252, RM400 and RM410), and for the detection of adulterant like Sharbati (RM215, RM423 and RM259). SSR database reported here, shall broaden the list of SSR markers already recommended for varietal identification and detection of adulteration in Basmati rice supplies.

Development of specific simple sequence repeat (SSR) markers for non-pollen type thermo-sensitive genic male sterile gene in rice (Oryza sativa L.)

A key factor of two-line hybrid rice production is the development of thermo-sensitive genic male sterility (TGMS) lines. In this study, a TGMS line showing non-pollen type thermo-sensitive genic male sterility was used. Crossing between TGMS line (female parent) and normal pollen varieties; CNT1 and PTT1 (male parent) were performed and F 1 and F 2 populations were develop for each crosses. The phenotypic segregation ratio, 3:1 (fertile: sterile pollens) observed in F 2 populations of TGMS/CNT1 and TGMS/PTT1 crosses confirmed that the non-pollen type thermo-sensitive genic male sterility is controlled by a single recessive gene. The bulk segregant analysis (BSA) using simple sequence repeat (SSR) markers were deployed to identify the location and genetic effect of this gene. We have generated new set of SSR markers to identify progenies carrying TGMS gene in a cross TGMS/PTT1. The TGMS gene was located on chromosome 2 with 0.0 cM distance from T2 marker. This TGMS gene was located in the same region of previous identified tmsX gene. It might be the same allele of this gene of 2 different lines. This marker could be used for marker assisted backcrossing to transfer TGMS gene for developing female parent in two line hybrid rice system. Key words: Hybrid rice, thermo-sensitive genic male-sterile, non-pollen type, molecular marker, marker assisted backcrossing.

Novel design and deployment of orthologous genic SSR markers in Eucalyptus camaldulensisDehnh

Background Eucalyptus camaldulensis is a widely planted tree species in India, because of its rapid growth and adaptability to dry regions. Genetic improvement through informed breeding of E. camaldulensis largely depends on availability of molecular markers, linkage maps and genome information. Microsatellite markers, also called as simple sequence repeats (SSRs) have wide application due to their unique advantages over other marker systems. In Eucalyptus, SSR markers have been used in various breeding applications from DNA fingerprinting to QTL mapping [1,4]. Despite of their advantages, the major drawback is the time required for their development [6]. There are less than twenty five E. camaldulensis specific SSR markers available in the public database [5]. Although, there are large number of SSRs available in other Eucalyptus species [2,3], their species transferability in camaldulensis is questionable for practical use. Nevertheless, the existing camaldulensis specific SSRs are insufficient for developing linkage maps, QTL and comparative mapping studies. Besides, EST based markers are handicapped due to exclusion of introns, which sometimes lead to compromise on product sizing. Therefore we have modeled a novel strategy that targets highly conserved domain in the genic region using both publicly available ESTs as well as genome sequences.

Molecular and biochemical studies of some yeast strains

This work focuses on detecting the level of polymorphisms among eleven KSA and Egyptian yeast strains, as efficient tools to assess the genetic relationships and development of yeast strain-specific molecular fingerprints. Moreover, estimation of the amino acid concentration was efficient in selecting the protein rich strains for animal feeding source. To detect the polymorphism among the yeast strains at the molecular level, 8 random amplified polymorphic DNA (RAPD), 5 inter simple sequence repeat (ISSR), and 8 specific simple sequence repeat (SSR) pair of primers were used. The total number of fragments produced by RAPD primers was 46 fragments and represented 52% of polymorphism. Also, number of fragments produced by ISSR and SSR primers was 45 fragments and recorded 63 and 77% of polymorphism among the strains, respectively. The amino acid analysis showed that yeast strains, Rhodotorula glutinis (Y.1); Schwanniomyces occidentalis (Y.2); Debaryomyces hansenii (Y.8); Kluyveromyces lactis (Y.9) and Pichia jadinii (Y.10) contained almost double the amount of total amino acids compared to the rest of the eleven strains used. The different molecular markers have confirmed each other and supported the biochemical analysis data, because the clustering analysis has shown that the previous five strains, (Y.1); (Y.2); (Y.8); (Y.9) and (Y.10), were falling together in the same sub cluster. Key words: Yeast, molecular markers, biochemical analysis.

EVALUATION OF SIMPLE SEQUENCE REPEATS (SSR) FOR GENETIC ANALYSIS OF CHESTNUT TREES IN MICHIGAN ORCHARDS

In Michigan, non-grafted seedlings of Chinese descent commonly planted by growers predominate chestnut orchards because of the resistance to chestnut blight. In addition, grafted hybrids of European, Chinese, and Japanese descent, many with unknown pedigrees, are also present in orchards. The outcome of this genetic mixing in orchards is a chestnut product with variable size and quality. Therefore, the objective of this study is to evaluate the use of simple sequence repeats (SSRs) to genetically characterize promising chestnut trees and cultivars currently growing in Michigan orchards. In the summer of 2007, we began a series of crosses in four isolated orchards, where each orchard consisted of two cultivars. Trees of the C. sativa × C. crenata hybrid cultivar 'Colossal' were selected as the female parent for every cross due to its good yield and because it is planted in orchards throughout the state. The chestnut cultivars 'Benton Harbor' and 'Eaton', both with Asian backgrounds, and the European hybrid cultivars 'Nevada', and 'Okei' were selected as pollinizer trees. Fifty nuts per cross were randomly selected as the individuals representing the four populations. We were able to generate fingerprints using both universal and specific primers such as (GTG) 5 and CsCAT/EMCs loci, respectively. The SSRs data were analyzed using 'PeakScanner' and 'Identity' software. Complex polymorphic bands were observed among chestnut cultivars, which ranged from 1,200 to 300 base pairs in length when (GTG) 5 was used. Amplicon sizes ranged from 76 to 193 base pairs when specific SSRs were used and the number of alleles observed at a locus ranged from 4 to 6. Results obtained from this study will allow for the genetic characterization of the chestnut cultivars and seedling trees currently growing in Michigan, and subsequently will help us establish high quality chestnut plantings for food and fiber production.

Development and Characterization of Large‐Scale Simple Sequence Repeats in Jute

ABSTRACTJute is an important crop of the Indian subcontinent and comprises tossa jute (Corchorus olitorius) and white jute (C. capsularis). The yield and fiber quality of this crop remained stagnant for many years and could not be improved through conventional plant breeding. Also, no effort has been made to develop molecular markers on a scale required for marker‐assisted selection (MAS) to supplement conventional plant breeding. As a first step toward deploying MAS for jute improvement, 2469 simple sequence repeats (SSRs) were developed in tossa jute (JRO 524) using four SSR‐enriched genomic libraries. A random subset of 100 SSRs (25 SSRs from each library) was used to detect polymorphism between the parental genotypes of each of the two recombinant inbred line (RIL) mapping populations. The RILs are being developed from JRO 524 × PPO4 (for fiber fineness) and JRC 321 × CMU 010 (for lignin content) crosses to prepare molecular maps and conduct quantitative trait loci (QTL) analyses. Both SSR length polymorphism and ± polymorphism (null alleles, i.e., presence and absence of specific SSR) were detected; 50 SSRs detected polymorphism between the two genotypes of tossa jute, whereas 45 SSRs detected polymorphism between the two genotypes of white jute. This SSR allelic polymorphism in jute is higher than that reported in other crops and is adequate for construction of genetic maps for QTL analysis. The large‐scale SSRs will also prove useful in studying genetic diversity, population structure, and association mapping.

Genetic analysis and localization of loci controlling leaf rust resistance of Triticum aestivum × Triticum timopheevii introgression lines

Introgressive lines resulting from crossing common wheat Triticum aestivum with the tetraploid T. timopheevii are characterized by effective resistance to leaf rust caused by Puccinia triticina Eriks. Molecular analysis using 350 specific simple sequence repeat (SSR) markers determined localization of the T. timopheevii genome in chromosomes 1A, 2A, 2B, 5A, 5B, and 6B. A population of F2 offspring of crossing hybrid line 842-2 with common wheat cultivar Skala was obtained for mapping the loci controlling leaf rust resistance. Analysis of association of phenotypic and genotypic data by means of simple interval mapping (SIM) and composite interval mapping (CIM) has shown that the resistance of adult plants is determined by two loci in chromosomes 5B and 2A. The major locus QLr.icg-5B, transferred from T. timopheevii chromosome 5G mapped to the interval of microsatellite loci Xgwm408-Xgwm1257 controls 72% of the phenotypic variance of the trait. The other, minor locus QLr.icg-2A located to chromosome 2A at a distance of 10 cM from Xgwm312 accounts for 7% of the trait expression. Microsatellite markers located near these loci may be used for controlling the transfer of agronomically valuable loci when new lines and cultivars are created.

Inter simple sequence repeat profile as a genetic marker system in sugarcane

Molecular profiling for varietal differentiation assumes an important role in the light of UPOV initiatives and PVP act. Inter simple sequence repeats (ISSR), a PCR based genetic marker system circumvents the requirement for flanking sequence information, and thus has found wide applicability in a variety of plants. The potential of ISSR markers for molecular profiling was assessed in sugarcane using a panel of 42 varieties of subtropical India. ISSR amplification was achieved using di-, tri-and tetraoligonucleotide repeats. All the nucleotide motifs were amenable to amplification. The ISSR results suggested that the frequency and clustering of specific simple sequence repeats was variable and motif-specific, however, there was not much difference in the di-, tri-or tetra-nucleotide ISSR profiles. The ISSR amplification pattern was used to assess genetic similarity of these sugarcane genotypes by Dice similarity coefficients, which ranged from 0.68 to 0.97 with a mean similarity value of 0.878. The ISSR amplification proved to be a valuable method for determining genetic variability among sugarcane varieties and for identification of the cultivars.

Molecular Mapping of the Russian Wheat Aphid Resistance Gene Dn2414 in Wheat

Russian wheat aphid (RWA) [Diuraphis noxia (Kurdjumov)] is an important pest of wheat (Triticum aestivum L.) in several production areas of the world. The most effective and economical approach for controlling RWA is to use resistant cultivars. A wheat line, ST‐ARS 02RWA2414‐11 (2414‐11), showed a high level of resistance to RWA biotype 2. Our objectives were to map the resistance gene and develop polymerase chain reaction (PCR)–based markers for marker‐assisted selection (MAS). A mapping population of 212 F2 individuals was developed from a cross of 2414‐11 and the susceptible cultivar Yuma. The F2 individuals and F2:3 families were infested using biotype 2 RWA. The RWA resistance of 2414‐11 is controlled by a single major gene, provisionally designated as Dn2414 Using standard PCR, 30 marker loci were found to be linked to Dn2414 with recombination frequencies (θ) of 0.00 to 0.27 and logarithm of the odds to the base 10 (LOD) scores of 7.6 to 66.1. Of the 30 markers, 26 were tightly linked to Dn2414 with θ ≤ 0.05. A genetic map was constructed consisting of 31 loci spanning a region of 34.7 cM. The close linkage of Dn2414 with several rye chromosome 1R short arm (1RS)‐specific simple sequence repeat markers and low θ values around the Dn2414 gene indicate that Dn2414 is located on chromosome 1RS.1BL (translocation chromosome with IRS and wheat chromosome 1B long arm). Phenotypic and marker profiles of 2414‐11 and its relatives are the same as other lines known to carry Dn7 The Dn2414 gene is thus located on 1RS arm, and the large number of PCR markers will be valuable for MAS of this gene.

An integrated molecular linkage map of diploid wheat based on a Triticum boeoticum × T. monococcum RIL population

Diploid A genome species of wheat harbour immense variability for biotic stresses and productivity traits, and these could be transferred efficiently to hexaploid wheat through marker assisted selection, provided the target genes are tagged at diploid level first. Here we report an integrated molecular linkage map of A genome diploid wheat based on 93 recombinant inbred lines (RILs) derived from Triticum boeoticum x Triticum monococcum inter sub-specific cross. The parental lines were analysed with 306 simple sequence repeat (SSR) and 194 RFLP markers, including 66 bin mapped ESTs. Out of 306 SSRs tested for polymorphism, 74 (24.2%) did not show amplification (null) in both the parents. Overall, 171 (73.7%) of the 232 remaining SSR and 98 (50.5%) of the 194 RFLP markers were polymorphic. Both A and D genome specific SSR markers showed similar transferability to A genome of diploid wheat species. The 176 polymorphic markers, that were assayed on a set of 93 RILs, yielded 188 polymorphic loci and 177 of these as well as two additional morphological traits mapped on seven linkage groups with a total map length of 1,262 cM, which is longer than most of the available A genome linkage maps in diploid and hexaploid wheat. About 58 loci showed distorted segregation with majority of these mapping on chromosome 2A(m). With a few exceptions, the position and order of the markers was similar to the ones in other maps of the wheat A genome. Chromosome 1A(m) of T. monococcum and T. boeoticum showed a small paracentric inversion relative to the A genome of hexaploid wheat. The described linkage map could be useful for gene tagging, marker assisted gene introgression from diploid into hexaploid wheat as well as for map based cloning of genes from diploid A genome species and orthologous genes from hexaploid wheat.

Rationalization of taro germplasm collections in the Pacific Island region using simple sequence repeat (SSR) markers

AbstractA regional (Oceania) core collection for taro germplasm has been developed based on phenotypic and molecular characterization. In total, 2199 accessions of taro germplasm have been collected byTaroGen(Taro Genetic Resources: Conservation and Utilisation) from 10 countries in Oceania: Papua New Guinea, Solomon Islands, Vanuatu, New Caledonia, Fiji, Palau, Niue, Tonga, Cook Islands and Samoa. Our objective was to select 10% from each country to contribute to a regional core. The larger collections from Papua New Guinea, Vanuatu and New Caledonia were analysed based on phenotypic characters, and a diverse subset representing 20% of these collections was fingerprinted. A diverse 20% subsample was also taken from the Solomon Islands. All accessions from the other six countries were fingerprinted. In total, 515 accessions were genotyped (23.4% overall) using taro specific simple sequence repeat (SSR) markers. DNA fingerprint data showed that great allelic diversity existed in Papua New Guinea and the Solomon Islands. Interestingly, rare alleles were identified in taros from the Solomon Islands province of Choiseul which were not observed in any of the other collections. Overall, 211 accessions were recommended for inclusion in the final regional core collection based on the phenotypic and molecular characterization.

Analysis of simple sequence repeats (SSRs) in wild barley from the Fertile Crescent: associations with ecology, geography and flowering time.

Wild barley, Hordeum spontaneum C. Koch, is the progenitor of cultivated barley, Hordeum vulgare. The centre of diversity is in the Fertile Crescent of the Near East, where wild barley grows in a wide range of conditions (temperature, water availability, day length, etc.). The genetic diversity of 39 wild barley genotypes collected from Israel, Turkey and Iran was studied with 33 SSRs of known map location. Analysis of molecular variance (AMOVA) was performed to partition the genetic variation present within from the variation between the three countries of origin. Using classification tree analysis, two (or three) specific SSRs were identified which could correctly classify most of the wild barley genotypes according to country of origin. Associations of SSR variation with flowering time and adaptation to site-of-origin ecology and geography were investigated by two contrasting statistical approaches, linear regression based on SSR length variation and linear regression based on SSR allele class differences. A number of SSRs were significantly associated with flowering time under four different growing regimes (short days, long days, unvernalised and vernalised). Most of the associations observed could be accounted for by close linkage of the SSR loci to earliness per se genes. No associations were found with photoperiodic and vernalisation response genes known to control flowering in cultivated barley suggesting that different genetic factors may be active in wild barley. Novel genomic regions controlling flowering time in wild barley were detected on chromosomes 1HS, 2HL, 3HS and 4HS. Associations of SSRs with site-of-origin ecological and geographic data were found primarily in genomic regions determining plant development. This study shows that the analyses of SSR variation by allele class and repeat length are complementary, and that some SSRs are not necessarily selectively neutral.

Development of simple sequence repeat markers in rye (Secale cerealeL.)

Simple sequence repeats (SSRs), also referred to as microsatellites, represent a PCR-based marker system that has been described in mammalian and plant genomes in recent years. In self-pollinating crop plants they have been shown to be superior to other DNA markers with respect to their level of polymorphism. The technical advantages compared with RFLP markers should also facilitate marker analysis in outcrossing crops like rye. In order to determine the usefulness of SSR markers in rye genetics and breeding, several genomic libraries were screened for (CT/GA)n and (GT/CA)n dinucleotide repeats. It was estimated that these motifs occur at a frequency of one per 268-519 kb. Seventy four out of 182 positive clones were sequenced, and the majority (56.8%) revealed perfect repeats, predominantly of the type (GT/CA)n (61.9%). Fifty seven primer pairs were designed and 27 (47.4%) resulted in specific SSR markers, of which 20 were genetically mapped or assigned to chromosomes or chromosome arms, respectively. The level of polymorphism of four SSR and three RFLP markers was assessed in two open-pollinated rye cultivars. On average, the SSR markers showed larger values of expected heterozygosity (0.62 vs. 0.43) and allele number (5.9 vs. 3.4) than RFLP markers in both cultivars.