Analyses of genetic diversity and population structure of anchote (Coccinia abyssinica (Lam.) Cogn.) using newly developed EST-SSR markers

Anchote (Coccinia abyssinica (Lam.) Cogn.) is a perennial root crop belonging to Cucurbitaceae family. It is endemic to Ethiopia and distributed over wide range of agro-ecologies. For further improvement and efficient conservation of this crop, characterization of its genetic diversity and its pattern of distribution is a vitally important step. Expressed sequence tags-simple sequence repeats (EST-SSRs) markers were developed from publicly available watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai] ESTs in the GenBank database. Among those novel markers, eight were polymorphic and subsequently used for genetic diversity and population structure analyses of 30 anchote accessions collected from western Ethiopia. A total of 24 alleles were obtained across the eight polymorphic loci and 30 accessions that revealed moderate level of genetic diversity in this minor crop. Among the eight loci, locus CA_06 was the most informative with six alleles and polymorphic information content (PIC) of 0.76. The accessions showed about threefold variation in terms of genetic diversity, with expected heterozygosity (He) ranging from 0.15 (accession An) to 0.44 (accession Dg). Other accessions with higher genetic diversity include Ar and Gu (He?=?0.43 and 0.41, respectively). Analysis of molecular variance (AMOVA) revealed that the variation within accessions and among accessions accounted for 84.7% and 15.3% of the total variation, respectively. The study revealed low but significant population differentiation in this crop with no clear pattern of population structure. The EST-SSR markers developed in this study are the first of their kind for anchote and can be used for characterization of its wider genetic resources for conservation and breeding purposes.

     

Genetic diversity in Tunisian perennial ryegrass revealed by ISSR markers

The genetic diversity in Tunisian perennial ryegrass (Lolium perenne) was examined by the help of inter-simple sequence repeats (ISSR). Starting from eighteen accessions, a large number of polymorphic ISSR markers were currently generated using appropriate primers (a total of 136, which average of 12.6 polymorphic bands/primer). These markers were considered to estimate the genetic distance among accessions and to draw phylogenetic trees. Our data provide evidence of a high degree of genetic diversity in Tunisian ryegrass. In addition, both cultivars and wild types present a high degree of divergence suggesting a complex domestication process in this crop. Moreover, spontaneous populations of Tunisian ryegrass have been identified as important ecotypes that are suitable in selection programs to improve grasslands.     

Genetic diversity of cultivated flax (Linum usitatissimum L.) and its wild progenitor pale flax (Linum bienne Mill.) as revealed by ISSR markers

Little is known about the genetic diversity of pale flax (Linum bienne Mill.), the wild progenitor of cultivated flax (L. usitatissimum L.), and ex situ germplasm of pale flax was scarce. Effort was made to collect 34 pale flax accessions and five landrace accessions of cultivated flax in Turkey. The inter simple sequence repeat (ISSR) technique was applied to characterize this set of flax germplasm, along with one Turkish cultivar, one Russian cultivar, five winter and four dehiscent type accessions of cultivated flax. Twenty-four ISSR primer pairs detected a total of 311 DNA fragments, of which 298 bands were polymorphic across 493 flax samples (roughly 10 samples per accession). These polymorphic bands had frequencies ranging from 0.002 to 0.998 and averaging 0.38. Accession-specific ISSR variation (Fst values) ranged from 0.469 to 0.514 and averaged 0.493. There was 49.3% ISSR variation resided among these 50 accessions, 35.9% harbored among landrace, winter, dehiscent types of cultivated flax and pale flax, and 38.2% present among 34 pale flax accessions. Pale flax displayed more ISSR variation than landraces and dehiscent type, but less than winter type, of cultivated flax. Clustering 493 individual plants revealed that these assayed plants were largely grouped according to their plant types and that pale flax was genetically more close to the dehiscent type, followed by the winter type and landrace, of cultivated flax. Pale flax collected within the geographic range of 180 km displayed a significant spatial genetic autocorrelation. Genetic distances among the pale flax accessions were significantly associated with their geographic distances and elevation differences. These findings are significant for understanding flax domestication and its primary gene pool.     

Genetic diversity of the African wild rice (Oryza longistaminata Chev. et Roehr) from Ethiopia as revealed by SSR markers

Data from microsatellite markers have been extensively used for both in situ and ex situ conservation strategies by determining the level of genetic diversity of natural populations that can widen the gene pool of cultivated plants. Such conservation practices are based on understanding of the between and within population genetic variations and partitioning populations on the basis of geographic origin. Therefore, the objective of this study was to assess the genetic diversity of Oryza longistaminata Chev. et Roehr and how this variation is partitioned within and between the eight O. longistaminata populations found in the different geographic regions of Ethiopia using simple sequence repeat markers. Five microsatellite markers in 320 samples generated 64 alleles that revealed the presence of large amount of genetic variability (Ho = 0.225; He = 0.768; Na = 7.375; Ne = 6.565 and P = 0.744). The F-statistics detected by the microsatellite loci showed Fst = 0.064 and Fis = 0.743 and there was no population in Hardy–Weinberg equilibrium. The genetic diversity results obtained from this data indicated that there are high levels of genetic diversity in the populations of O. longistaminata studied and it is higher within than between populations. Among the eight populations sampled, five populations were identified as priorities for conservation strategies. Thus, national collection and conservation strategies need to consider these populations.     

Genetic diversity and systematic relationships in sweetpotato (Ipomoea batatas (L.) Lam.) and related species as revealed by RAPD analysis

Summary Fifteen 10mer primers, in combination with the Stoffel fragment, were used to detect random amplified polymorphic DNA (RAPDs) among 26 accessions of sweetpotato (I. batatas (L.) Lam.) from Oceania, Peru, the Philippines, and the United States and between 8 Ipomoea species from section Batatas. Phenetic and principal coordinate analysis of the 56 polymorphisms detected within the hexaploid I. batatas clearly delineated the South Pacific and the Peruvian sweetpotato lines. The two U.S. cultivars clustered with the Oceanic materials. Cladistic and phenetic analysis of 8 Ipomoea species supports previously published phylogenies based on morphological and RFLP data. Among the species examined, I. tabascana, I. trifida and the tetraploid forms of I. batatas from Mexico and Ecuador, including I. batatas var. apiculata, are the taxa most closely related to the cultivated hexaploid I. batatas. These findings support the utility of RAPD markers for evaluating genetic diversity in sweetpotato and for establishing taxonomic and evolutionary relationships in Ipomoea.     

Genetic diversity in cowpea [Vigna unguiculata (L.) Walp.] as revealed by RAPD markers

The present study, using RAPD analysis, was undertaken to characterize genetic variation in domesticated cowpea and its wild progenitor, as well as their relationships. The materials used consisted of 26 domesticated accessions, including accessions from each of the five cultivar-group, and 30 wild/weedy accessions, including accessions from West, East and southern Africa. A total of 28 primers generated 202 RAPD bands. One hundred and eight bands were polymorphic among the domesticated compared to 181 among wild/weedy cowpea accessions. Wild accessions were more diverse in East Africa, which is the likely area of origin of V. unguiculata var. spontanea. Var. spontanea is supposed to have spread westward and southward, with a loss of variability, loss counterbalanceed in southern Africa by introgressions with local perennial subspecies. Although the variabilty of domesticated cowpea was the highest ever recorded, cultivar-groups were poorly resolved, and several results obtained with isozyme data were not confirmed here. However primitive cultivars were more diverse than evolved cultivars, which still suggests two consecutive bottlenecks within domesticated cowpea evolution. As isozymes and AFLP markers, although with a larger number of markers, RAPD data confirmed the single domestication hypothesis, the gap between wild and domesticated cowpea, and the widespread introgression phenomena between wild and domesticated cowpea.     

Genetic variability of wild apricot (Prunus armeniaca L.) populations in the Ili Valley as revealed by ISSR markers

The genetic variability of 14 wild Prunus armeniaca populations was investigated using morphological analysis and inter-simple sequence repeat markers. 10 morphological characters revealed a high level variation, especially Fruit number, Fruit weight, Seed weight and Tree height. Totally, 15 selected primers generated 155 loci, with an average of 10.3 bands per primer. Nei’s gene diversity (H _e ) and Shannon’s index of diversity (I) were fairly high at the species level (H _e  = 0.2741, I = 0.4220). High molecular and morphological variability indicated that wild apricots in the Ili Valley still maintained a relatively high level of diversity. The G _ST of 0.2275 revealed a low level of genetic differentiation among populations, and genetic variation mainly resided within populations (81.51 %), which was identified with the moderate gene flow value (N _m  = 1.6974). The relatively high intraspecific genetic diversity and low inter-population genetic differentiation was largely attributed to long-distance dispersal of pollen, continuous distribution of populations and the self-incompatible breeding system.     

Genetic diversity of Pakistan wheat germplasm as revealed by RAPD markers

Wheat breeding in Pakistan started in 1930s before partition in the United India and so far has released more than 68 cultivars, but no systematic analyses of the genetic diversity of Pakistan wheat have been made. Twenty Pakistan wheat cultivars released from 1933 to 2002 were examined for genetic diversity and relationships using random amplified polymorphic DNA (RAPD) markers. Forty-two RAPD primers were applied and 184 polymorphic bands were generated for each cultivar. Most of the cultivars were genetically interrelated, although six of them displayed some genetic distinctness. The RAPD variation observed among these cultivars was low. Only 40.7% of the total scorable bands were polymorphic, and 26.1% of the polymorphic bands were observed most frequently (f = 0.95) among the 20 cultivars. The proportions of polymorphic bands for each cultivar ranged from 0.67 in ‘Yecora’ to 0.84 in ‘C-250’ with an average of 0.76. About 1.4% of the RAPD variation might have been fixed over the 69 years of wheat breeding, but such fixation was not statistically significant. These results are significant for future improvement and conservation of Pakistan wheat.     

Genetic diversity among Northwestern Argentinian cultivars of common bean (Phaseolus vulgaris L.) as revealed by RAPD markers

The genetic diversity of 10 commercial cultivars of common beans, developed in Northern Argentina was analyzed based on RAPD markers. Sixteen primers were assayed and among them only 4 showed polymorphisms. A similarity matrix was generated by applying three different association coefficients, Simple Matching, Jaccard and Dice. By the UPGMA method dendrograms were generated and also the principal coordinate analysis was performed. The similarity values found were higher than 40% suggesting that genetic diversity is low. Both cluster analysis and principal coordinates analysis associated commercial cultivars either to the Andean or the Mesoamerican gene pool.     

Genetic diversity and population structure of vegetable soybean (Glycine max (L.) Merr.) in China as revealed by SSR markers

Vegetable soybean is a kind of value-added specialty soybean serving as vegetable or snacks. Understanding the genetic structure of vegetable soybean is a key point for further utilization in breeding programs. In the present study, the genetic structure and diversity of 100 vegetable soybean accessions planted in China was analyzed using 53 simple sequence repeat (SSR) markers. A total of 296 alleles were detected with an average of 5.6 alleles per SSR locus. The polymorphism information content (PIC) values of SSR markers ranged from 0.074 to 0.831, with an average of 0.573. Nei’s genetic distance between accessions ranged from 0 to 0.9434 with an average of 0.6286. These vegetable soybean germplasms could be divided into 8 subgroups based on STRUCTURE analysis, or 11 subgroups based on unweighted pair group method with arithmetic average (UPGMA) cluster. Further comparison showed that the UPGMA subgroups and STRUCTURE subgroups were in fact highly consistent. Germplasms in each classified groups showed great consistency with their origins, seed coat colors or pedigrees. Genetic relationships among germplasm panels that initially came from different geographical regions were also analyzed. Germplasm panels from China Mainland, Taiwan Island and Japan were highly similar to each other with the similarities of over 98 %. Molecular data and cluster analysis also showed that germplasms from China Mainland are more diverse than those from other areas. These results gave us a deep insight into the genetic structure of vegetable soybeans in China and will help us to improve the breeding strategies.     

Genetic diversity in Japanese aromatic rice (Oryza sativa L.) as revealed by nuclear and organelle DNA markers

Aromatic rice (Oryza sativa L.) cultivated in Japan is regionally differentiated by geographical distribution and characteristics. We aimed to characterize the lineage of Japanese aromatic rice using DNA markers. Based on analyses with nuclear SSR markers, we found that Japanese aromatic rice cultivars belong, with one exception, to japonica but showed some differences from authentic japonica and were divided into two clades that were distributed in western and eastern Japan, respectively. Further analyses with organelle markers showed that most of the cultivars in eastern Japan had cytoplasm characterized by tropical japonica, whereas most of those in western Japan had cytoplasm characterized by temperate japonica. We postulate that the ancestor of the cultivars in eastern Japan differs from those of the cultivars in western Japan, and that the two groups may have been separately introduced from Taiwan into Japan. The cytoplasm of aromatic rice cultivars in western Japan may have originated from tropical japonica and been substituted into the cytoplasm of temperate japonica through hybridization between tropical japonica as a male parent and temperate japonica as a female parent.     

Genetic diversity in spring faba bean (Vicia faba L.) genotypes as revealed by high-throughput KASP SNP markers

Genetic Resources and Crop Evolution - Faba bean is one of the oldest legume crop grown around the world for its cheap source of protein and for being used in crop rotation with cereals to improve...     

Genetic diversity analysis of yams (Dioscorea spp.) cultivated in China using ISSR and SRAP markers

Yam (Dioscorea spp.) is widely cultivated in China and many landraces are maintained by local farmers. However, there is little information available about their diversity and species identity. In this study, inter simple sequence repeat (ISSR) and sequence related amplified polymorphism (SRAP) techniques were used to assess genetic diversity within 21 yam landraces from seven cultivated populations. We observed high level of polymorphism among these landraces, specifically, 95.3 % for ISSR and 93.5 % for SRAP. Analysis of molecular variance revealed a significantly greater variation among the four yam species (40.39 %) and their populations (35.78 %) than within the populations (23.83 %). The unweighted pair group method arithmetic averages clusters and principal component analysis for 21 landraces formed four well-separated groups containing landraces of each of the four species, namely, Dioscorea opposita Thunb., Dioscorea alata L., Dioscorea persimilis Prain et Burkill, and Dioscorea fordii Prain et Burkill. The ISSR and SRAP primers were highly discriminatory among the 21 landraces; all 21 landraces could be easily differentiated using these primers. The average mean of gene flow (Nm = 0.1081) estimated from high genetic differentiation (Gst = 0.8222) suggested that gene flow among the populations was relatively restricted. The lack of genetic diversity within individual yam species suggests that it is critical to develop long-term strategies for enhancing genetic diversity within various yam species.     

Genetic diversity and relationships among cultivated and wild accessions of tartary buckwheat (Fagopyrum tataricum Gaertn.) as revealed by RAPD markers

The methodology of sampling and the selection of a proper marker systemfor the analysis of accessions are major concerns in the evaluation of gene bank material. In our study the RAPD analysis of bulked DNA samples and single seedsDNA was successfully employed to evaluate intra- and inter-population geneticvariability of cultivated and wild tartary buckwheat accessions. The bulkingapproach enabled the distinction of all 40 analysed accessions and theirseparation into geographically well defined clusters. Three wild populations,two from Sichuan and one from Qinghai, formed a group that was geneticallyrelatively distant from wild populations from Tibet and all cultivatedlandraces which, on the other hand, exhibited very close relationships. Thesingle seed study that was used after bulked DNA analysis provided detailedinformation of the genetic variation present within some accessions of specialinterest. A moderate level of genetic variability was detected betweenaccessions and the variability was partitioned into between- andwithin-population components. On average, most of the detected variation ispresent between F. tataricumpopulations. The genetic and geographic distribution of variability is furtherdiscussed. We demonstrated the usefulness of combining bulking and single seedstudy approaches for the effective evaluation of genetic variability inF. tataricum accessions that couldalso have wider applicability in the management of plant genetic resources andphylogenetic studies.     

Genetic diversity in Nordic horseradish,Armoracia rusticana,as revealed by AFLP markers

176 horseradish accessions from the Nordic countries Denmark, Finland, Norway and Sweden were analysed to estimate the level of genetic diversity and to propose conservation strategies for this cultivated plant. Most of the accessions were collected in old gardens of the Nordic countries but selections from European countries and Danish breeding lines were also included in the study. Since horseradish is mainly vegetative propagated the genetic diversity has been assumed to be small. However, using the AFLP method with three primer combinations we revealed a significant genetic diversity among Nordic horseradish. The analysis yielded 65 polymorphic bands and we found an overall diversity index of 0.5 (Shannon–Weaver). The highest diversity was found among the Finnish accessions followed by the Danish accessions. An overall AMOVA analysis indicated that 90 % of the variation could be explained by among accession variation. The AFLP data assigned the different accessions into groups that corresponded with their country of origin. A closer relationship was observed between the Swedish, Danish and some of the Norwegian accessions while the Finnish accessions separated more clearly from the other three countries. A possible explanation for the diversity is that horseradish probably has been introduced to the Nordic countries at many occasions during a long period of time.     

Genetic diversity in sweetpotato [Ipomoea batatas (L.) Lam.] in relationship to geographic sources as assessed with RAPD markers

Available evidence shows that sweetpotato originated from either Central or South American lowlands with subsequent dispersal to North America, Europe, Africa, Asia and the pacific islands. A total of 71 polymorphic RAPD molecular markers were used to assess the genetic relationships amongst 74 sweetpotato varieties originating from a total of 23 sweetpotato producing countries within six geographical regions, namely, South America, Central America/Caribbean, United States of America (USA), East Africa, Asia and Oceania. An Analysis of Molecular Variance (AMOVA) indicated that 93.4% of the total variance was due to the differences between genotypes within regions. The difference between regions was significant (P < 0.001) but only contributed 6.6% to the variance. Genetic distance (PhiST) calculated with AMOVA and multidimensional scaling (MDS) revealed that the South American and the Central American/Caribbean genotypes formed two separate clusters. East African varieties, which have unique characteristics from other traditional varieties, were distinct from other traditional varieties from South America and Oceania. These results support the reported hypothesis of the origin and dispersal of the sweetpotato and indicate that the primary centre of diversity probably has two distinct genepools. It is proposed that the dispersal of the sweetpotato from its origin may have mainly involved varieties from Central America/Caribbean as opposed to varieties from South America. There is an indication that new genepools may be evolving in Africa and Asia due to hybridisation and adaptation to the local environments.     

Genetic diversity and population structure of masson pine (Pinus massoniana Lamb.) superior clones in South China as revealed by EST-SSR markers

Genetic Resources and Crop Evolution - Masson pine (Pinus massoniana Lamb.) is one of the most economically important and widely distributed forest species in China. Understanding the genetic...     

Genetic diversity and population structure of Guinea yams and their wild relatives in South and South West Ethiopia as revealed by microsatellite markers

Genetic diversity and population structure of Guinea yams and their wild relatives collected from south and south west Ethiopia were assessed using microsatellite markers. The total number of alleles amplified for the 7 loci studied was found to be 60, with an average of 8.6 alleles per locus. The average expected heterozygosity for the entire population was found to be 64 % indicating that Guinea yams and their wild relatives in the study area display a high level of genetic diversity. Using allelic richness as a measure of genetic diversity the wild forms exhibited greater allelic diversity than the cultigens. Contrary to what is expected in vegetatively propagated crops, none of the seven loci studied showed a significant excess of heterozygotes. In all the comparisons made, a low mean F_ST (but significant) has been observed, indicating that the majority of microsatellite diversity in the populations under study was found within rather than between populations.     

Genetic diversity and parentage in farmer varieties of cacao (Theobroma cacao L.) from Honduras and Nicaragua as revealed by single nucleotide polymorphism (SNP) markers

Cacao (Theobroma cacao L.) is the main source for chocolate with an annual production of four million tons worldwide. This Neotropical tree crop was domesticated in Mesoamerica as far back as 3,000 years ago. Knowledge of genetic diversity and population structure in farmer varieties of cacao in the center of domestication is essential for sustainable production of fine-flavored cacao beans and contributes to in situ/on-farm conservation of farmer varieties. Based on 70 single nucleotide polymorphism markers, we analyzed 84 fine-flavored farmer varieties collected from traditional cacao farms in Honduras and Nicaragua. The study also included 31 clones from the international cacao collections to serve as references. The SNP based multilocus matching identified six synonymous groups, including 14 Criollo and two Amelonado varieties. A moderately high level of genetic diversity was observed in these farmer varieties, indicating the possibility to further explore intra-population variation and breed for fine-flavored cocoa. Multivariate analysis showed clustering of the 84 farmer accessions in five genetic groups: ancient Criollo, Amelonado, Trinitario (including Nicaragua Trinitario and Honduras Trinitario) and Upper Amazon Forastero (only one accession). The Honduras Trinitario differed from the Nicaragua Trinitario group. The clustering results largely supported the perceived classification of cacao by local farmers and researchers, which was mainly based on morphological traits. However, the well known traditional variety “Indio” in this region was identified as synonymous with Amelonado. Parentage analysis showed that the variety “Indio” (or Amelonado) contributed more to the Trinitario type farmer varieties, whereas ancient Criollo had less influence. The present study demonstrates the efficacy of using a small set of SNP makers for cacao germplasm characterization, and further depicts the diverse origins and parentage in farmer varieties from Mesoamerica. This information thus will be highly useful for conservation and utilization of cacao germplasm from this region.