Genetic variation of a global germplasm collection of chickpea (<Emphasis Type="Italic">Cicer arietinum</Emphasis> L.) including Italian accessions at risk of genetic erosion

Chickpea (Cicer arietinum L.) is one of the most important legumes worldwide. We addressed this study to the genetic characterization of a germplasm collection from main chickpea growing countries. Several Italian traditional landraces at risk of genetic erosion were included in the analysis. Twenty-two simple sequence repeat (SSR) markers, widely used to explore genetic variation in plants, were selected and yielded 218 different alleles. Structure analysis and hierarchical clustering indicated that a model with three distinct subpopulations best fits the data. The composition of two subpopulations, named K1 and K2, broadly reflects the commercial classification of chickpea in the two types desi and kabuli, respectively. The third subpopulation (K3) is composed by both desi and kabuli genotypes. Italian accessions group both in K2 and K3. Interestingly, this study highlights genetic distance between desi genotypes cultivated in Asia and Ethiopia, which respectively represent the chickpea primary and the secondary centres of diversity. Moreover, European desi are closer to the Ethiopian gene pool. Overall, this study will be of importance for chickpea conservation genetics and breeding, which is limited by the poor characterization of germplasm collection.     

Study on chloroplast DNA diversity of cultivated and wild pears (<Emphasis Type="Italic">Pyrus</Emphasis> L.) in Northern China

Eight pairs of chloroplast DNA (cpDNA) universal primers selected from 34 pairs were used to assess the genetic diversity of 132 pear accessions in Northern China. Among them, six amplified cpDNA fragments showed genetic diversity. A total of 24 variable sites, including 1 singleton variable site and 23 parsimony informative sites, as well as 21 insertion-deletion fragments, were obtained from the combined cpDNA sequences (5309–5535 bp). Two trnL-trnF-487 haplotypes, five trnL-trnF-413 haplotypes, five rbcL haplotypes, six trnS-psbC haplotypes, eight accD-psaI haplotypes and 12 rps16-trnQ haplotypes were identified among the individuals. Twenty-one haplotypes were identified based on the combined fragments. The values of nucleotide diversity (P_i), average number of nucleotide differences (k) and haplotype diversity (H_d) were 0.00070, 3.56408 and 0.7960, respectively. No statistical significance was detected in Tajima’s D test. Remarkably, the important cpDNA haplotypes and their representing accessions were identified clearly in this study. H_19 was considered as one of the ancient haplotypes and was a divergent centre. H_16 was the most common haplotype of the wild accessions. H_2 was the haplotype representing the most pear germplasm resources (46 cultivars and two wild Ussurian Pear accessions), followed by haplotype H_5 (30 cultivars, two wild Ussurian Pear accessions and four sand pears in outgroups) representing the cultivars ‘Dangshan Suli’ and ‘Yali’, which harbour the largest and the second largest cultivation areas in China. More importantly, this study demonstrated, for the first time, the supposed evolution routes of Pyrus based on cpDNA divergence in the background of pear phylogeny in Northern China.     

Genetic structure of cultivated <Emphasis Type="Italic">Zanthoxylum</Emphasis> species investigated with SSR markers

Zanthoxylum is an economically and ecologically important genus of the Rutaceae family, of which Z. bungeanum and Z. armatum have a long history of cultivation in China. However, how the natural processes such as selection and drift and agriculture practices have influenced the genetic variation of cultivated Zanthoxylum species during long-term domestication remains elusive. Herein, we determined the population genetic structure of current widely cultivated Zanthoxylum species, Z. bungeanum and Z. armatum. Microsatellite markers revealed a high level of genetic variation and significant genetic differentiation for both species despite Z. bungeanum showed higher genetic diversity than Z. armatum. AMOVA indicated that most of the genetic variation exists within individuals rather than among provenances for both species. Population structure analyses generated three distinct groups within the entire accessions. All Z. bungeanum accessions were distinguished into two major geographic groups, north and south groups, with Qinling Mountains as the main geographic barrier to gene flow while a significant genetic differentiation was observed between cultivated and wild Z. armatum accessions. Mantel test of Z. bungeanum displayed a significant correlation between genetic and geographic distances within each inferred group but no correlation between genetic and geographic distance was observed when comparing genetic and geographic distances focusing only on pairwise of north vs. south provenances, ruling out the hypothesis that gene flow between north and south provenances followed an isolation-by-distance model. Our research provided a fundamental genetic profile that will improve the conservation and responsible exploitation of the extant germplasm of Zanthoxylum.     

Analysis of genetic diversity and population structure in <Emphasis Type="Italic">Capsicum</Emphasis> landraces from North Eastern India using TE-AFLP markers

North eastern (NE) India harbours a precious germplasm repository of Capsicum in the form of various landraces. The present study was undertaken to characterise the extent of genetic variation present in different Capsicum landraces from north eastern India. A set of 171 Capsicum accessions were characterised using three-endonuclease amplified fragment length polymorphism (AFLP) markers. Out of 416 bands obtained from six primer combinations, 254 (61 %) were polymorphic. The pairwise genetic dissimilarity among accessions ranged from 0.03 to 0.97. Cluster analysis based on neighbour joining showed two major clusters. Cluster I contained most of the bhut jolokia accessions whereas cluster II contained all of the Capsicum annuum genotypes. Similar grouping was observed with population STRUCTURE analysis as well as principle coordinate analysis. Analysis of molecular variance (AMOVA) revealed 45 and 54 % variation among and within populations, respectively. This information on population structure analysis and molecular characterisation will be helpful for effective utilisation of this germplasm in Capsicum improvement programs.     

Development of cost-effective single nucleotide polymorphism marker assays for genetic diversity analysis in <Emphasis Type="Italic">Brassica rapa</Emphasis>

Competitive allele-specific PCR (KASPar) assay is a user-friendly system that provides flexibility in the numbers of single nucleotide polymorphisms (SNPs) and genotypes. Based on Illumina-GA-IIx genomic data from 10 genotypes with a broad genetic background, 3183 SNPs were selected for KASPar assays development, and 568 were finally converted and selected for Brassica rapa germplasm characterization (17.8%) on the basis of reproducibility, missing data rate, and uniform genetic distribution. High levels of polymorphism of these markers across 231 B. rapa genotypes were verified, illustrating by high polymorphic information content (averaged 0.34), minor allele frequency (0.37), genetic diversity (0.45), and the low observed heterozygosity (0.10). Based on the SNP dataset, structure and principal coordinates analysis, and neighbor-joining phylogenetic methods were used to examine the population structure and gave highly consistent results. The 231 accessions were divided into the four primary subspecies, representing 99 accessions from B. rapa ssp. pekinensis, 85 from B. rapa ssp. chinensis, 30 from B. rapa ssp. rapifera, and 17 from B. rapa ssp. oleifera and were further subdivided into 12 lower-order clusters according to different morphotypes. The genetic variability and pairwise fixation index analysis revealed that the ssp. pekinensis accessions possess the most extensive genetic variation among the four subspecies. The KASPar system is highly useful for validating SNPs and will be valuable for genetics research and breeding applications in B. rapa.     

<Emphasis Type="Italic">Miscanthus</Emphasis> sp.: Genetic Diversity and Phylogeny in China

Miscanthus genetic resources are widely distributed throughout China. However, genetic studies on Miscanthus lagged far behind other crops (e.g., sorghum, maize). To establish the comprehensive genetics knowledge of Miscnathus in China, here we report the genetic and phylogenetic diversity of 174 domestic Miscanthus accessions, along with an external Miscanthus × giganteus control. Cytological observations and flow cytometry analyses indicated that there were two major Miscanthus cytotypes in China: diploid (86.86%) and tetraploid (12.57%) without triploid. A total of 108 polymorphic loci generated from 25 SSR primers were used to evaluate the genetic variation. Large variations in genetic similarity coefficients (GSCs), ranging from 0.08 to 0.97 with a mean value of 0.39, were observed between these Miscanthus accessions. Our phylogenetic data revealed that these accessions were clustered into four main clades: M. section Miscanthus, M. section Diandranthus, M. section Triarrhena, and hybrids. The average percentage of polymorphic loci (P), gene diversity (H), and Shannon’s diversity index (I) among Miscanthus species are 70.93%, 0.22, and 0.34, respectively. These were consistent with the analysis of molecular variance (AMOVA) results, showing that 85% of genetic variation was found within clades. This study investigated the clear phylogenetic relationship of Miscanthus species in China, which will be valuable for further utilization of the germplasm in genetic improvement and hybrid breeding of Miscanthus.     

Evidence of the genetic diversity and clonal population structure of <Emphasis Type="Italic">Oenococcus oeni</Emphasis> strains isolated from different wine-making regions of China

Studies of the genetic diversity and population structure of Oenococcus oeni (O. oeni) strains from China are lacking compared to other countries and regions. In this study, amplified fragment length polymorphism (AFLP) and multilocus sequence typing (MLST) methods were used to investigate the genetic diversity and regional evolutionary patterns of 38 O. oeni strains isolated from different wine-making regions in China. The results indicated that AFLP was markedly more efficient than MLST for typing O. oeni strains. AFLP distinguished 37 DNA patterns compared to 7 sequence types identified using MLST, corresponding to discriminatory indices of 0.999 and 0.602, respectively. The AFLP results revealed a high level of genetic diversity among the O. oeni strains from different regions of China, since two subpopulations and an intraspecific homology higher than 60% were observed. Phylogenetic analysis of the O. oeni strains using the MLST method also identified two major phylogroups, which were differentiated into two distinct clonal complexes by minimum spanning tree analysis. Neither intragenic nor intergenic recombination verified the existence of the clonal population structure of the O. oeni strains.     

Assessment of genetic diversity in Ethiopian cowpea [<Emphasis Type="Italic">Vigna unguiculata</Emphasis> (L.) Walp.] germplasm using simple sequence repeat markers

The genetic diversity of cowpea (Vigna unguiculata L. Walp.) in Ethiopia was analyzed using 19 uniform accessions, 62 variable accessions (yielding 185 sub-types), and two mungbean (Vigna radiata) accessions (four subtypes) as outgroup. A set of 23 polymorphic simple sequence repeat (SSR) markers was identified, and polymorphism in the various accessions was scored by determining amplicon variability. Allele frequency, genetic diversity, and polymorphism information content (PIC) were determined for each SSR marker, and a neighbor joining dendrogram was generated to show the genetic relationship among the individual accessions. A total of 75 allelic variants was defined, with the average number of alleles per locus calculated to be three. The average genetic diversity (D) was 0.47, and PIC was 0.4. Three main clusters were identified by phylogenetic analysis, and the clusters and sub-grouping were supported by STRUCTURE and principal component analysis. This grouping had a moderate fixation index value of 0.075 and gene flow (Nm) of 3.176, indicating that the accessions possess wide diversity within individuals and populations. The accessions showed no clustering by geographical origins. Three well-characterized molecular markers (SSR1, C42-2B, and 61RM2) for race specific resistance to Striga gesnerioides in the cowpea cultivar B301 were used to evaluate the accessions for their potential for use in genetic improvement against this pest. Based on this analysis, only two accessions, 222890–2 from Gambela and 286–2 from the Southern Nations, Nationalities, and Peoples (SNNP) region, were found to cluster with B301 and contain the SSR1 resistance allele. These findings will assist in germplasm conservation efforts by the Institute of Biodiversity and Conservation of Ethiopia, and contribute to future studies aimed at the genetic improvement of local germplasm for improved overall agronomic performance as well as Striga resistance in particular.     

Genetic diversity and population structure of common walnut (<Emphasis Type="Italic">Juglans regia</Emphasis>) in China based on EST-SSRs and the nuclear gene phenylalanine ammonia-lyase (<Emphasis Type="Italic">PAL</Emphasis>)

Common walnut (Juglans regia L.) is an economically important temperate tree species valued for both its nut and wood. We investigated the genetic diversity and population structure of J. regia germplasm from 13 locations in China using 10 markers derived from expressed sequences (EST-SSR) and sequence polymorphisms within the phenylalanine ammonia-lyase (PAL) gene. Analysis of the population genetic structure based on EST-SSRs showed distinct populations in northern versus southern China that were not reflected in the spatial distribution of PAL haplotypes. High levels of population differentiation were probably the result of reproductive isolation and in southern China, hybridization with Juglans sigillata. The results indicate the possible presence of distinct evolutionary lineages of J. regia in the Qinling and Daba Mountains of China and in Yunnan province that may require ecological management if they are to be retained as in situ resources.     

The southwestern origin and eastward dispersal of pear (<Emphasis Type="Italic">Pyrus pyrifolia</Emphasis>) in East Asia revealed by comprehensive genetic structure analysis with SSR markers

Pyrus pyrifolia is considered one of the most important cultivated Pyrus species. Hundreds of landraces and bred cultivars have been developed through the natural and artificial hybridizations necessary due to self-incompatibility. In this study, the genetic diversity of 478 Pyrus accessions, including Chinese landraces, bred cultivars, and wild samples, as well as introduced pear cultivars from Japan and Korea, was investigated with a set of 17 simple sequence repeat (SSR) markers distributed across all 17 linkage groups of the pear genome. A total of 121 alleles were detected, including 4 rare alleles with a frequency lower than 5%. Diversity statistics indicated a high level of genetic variation as quantified by the average values of the observed heterozygosity, the expected heterozygosity, and Wright’s fixation index, at 0.76, 0.78, and 0.02, respectively. Population structure and discriminant analysis of principal component analysis implied extensive genetic communication between sand pears in China and revealed four contiguous geographical clusters with overlapping geographical regions. The diversity of the four clusters and approximate Bayesian computation (ABC) indicated that sand pear spread from west to east along the Pearl River and Yangtze River valleys. High diversity and polyphyletic genetic components of cultivars in southwestern China further support southwestern China as the probable center of divergence for Pyrus species. A core collection of 80 out of 470 cultivars was selected, accounting for about 17% of accessions, and capturing 91% of all alleles, including all rare alleles. Our research provides a comprehensive understanding of sand pear germplasm in East Asia and constructs a preliminary core collection, which will be useful for association genetics studies, germplasm conservation, and breeding programs.     

Genetic Diversity,Genotype Discrimination,and Population Structure of Mexican <Emphasis Type="Italic">Opuntia</Emphasis> sp., Determined by SSR Markers

The Opuntia (prickly pear) genus, an important horticultural crop in Mexico, is essentially a fruit crop with two variants: sweet (“tunas”) or acid (“xoconostles”) fruits; it is also a source of vegetables “nopalitos” or fodder for livestock, among other uses. Its taxonomical classification has been reported as complex, although few studies on the genetic structure of Mexican Opuntia are available, and genetic differences between the two types of fruits are unknown. Opuntia genotype identification and classification are still mainly based on morphological characters. In this study, the genetic diversity of Mexican Opuntia germplasm with agronomic and economic importance was revealed, using 88 accessions and 13 SSR markers, in an attempt to explore the genetic relationships among them. A total of 159 alleles were detected ranging from 7 to 23 per locus with an average of 12.2. The SSR markers generated unique fingerprints for each Opuntia accession confirming their usefulness for genetic analysis. The accessions’ grouping was defined by several complementary clustering methods, and the moderate incongruences between the different methods did not influence the overall clustering. DAPC and STRUCTURE analyses grouped the accessions into five groups, thus confirming the incorrect delimitation of species in this genus. The following species had no clear boundaries: Opuntia ficus-indica, Opuntia albicarpa, Opuntia megacantha, Opuntia streptacantha, Opuntia lasiacantha, and Opuntia hyptiacantha. However, Opuntia robusta was separated from the rest of the species. Opuntia joconostle and Opuntia matudae, which produce acid fruits, tended to differ from the others. Median-joining simulation classified all genotypes into a complex network, and both linear and reticular ties between Mexican Opuntia genotypes were revealed. The genetic distance revealed in the present study shows the importance of Mexican accessions for conservation and use in breeding programs.     

An interstate highway affects gene flow in a top reptilian predator (<Emphasis Type="Italic">Crotalus atrox</Emphasis>) of the Sonoran Desert

Roads can substantially impact the population connectivity of a wide range of terrestrial vertebrates, often resulting in loss of genetic diversity and an increase of spatial genetic structure. We studied the Western Diamond-backed Rattlesnake (Crotalus atrox), a large and abundant venomous predator, to test the hypothesis that a large and relatively new roadway in Arizona (Interstate Highway I-10) is a barrier that impacts gene flow and population genetics via habitat fragmentation. Based on 72 C. atrox sampled from three specific sampling sites (“subpopulations”) on both the west and east corridors of I-10, we used 30 nuclear microsatellite DNA loci and three mitochondrial DNA genes (2615 bp) to assess genetic diversity and structure, estimate effective population size (N _e ), and describe patterns of gene flow. We found no evidence for loss of genetic diversity or a decrease in N _e between the three subpopulations. Our microsatellite analysis showed that two subpopulations in close proximity (4 km), but separated by I-10, showed greater levels of genetic differentiation than two subpopulations that were separated by a greater distance (7 km) and not by I-10 or any other obvious barriers. Mitochondrial DNA analyses showed no significant genetic differentiation nor any indication of historically impeded gene flow. Tajima’s D and mismatch distribution tests revealed that demographic expansion is occurring in the overall population (all three subpopulations). Bayesian clustering and spatial genetic autocorrelation analyses of microsatellite data showed resistance to gene flow at the approximate location of I-10. Simulations that investigated gene flow between the subpopulations (with and without a highway barrier present) were consistent with our molecular results. We conclude that I-10 has reduced gene flow in a population of an important reptilian predator of the Sonoran Desert in southern Arizona and make conservation recommendations for reversing this trend.     

Assessment of genetic diversity among okra genotypes using SSR markers

Okra (Abelmoschus esculentus) is an important nutritious vegetable. Despite its high economic and industrial value, very little attention has been paid to assess genetic diversity of okra at molecular level. For effective conservation and proper deployment of germplasm, a study on diversity analysis of okra germplasm was conducted with DNA markers. Microsatellite/Simple sequence repeat (SSR) markers were utilized to evaluate the genetic diversity among 96 accessions of Abelmoschus, of which 92 accessions were of A. esculentus and one accession each of A. tuberculatus, A. moschatus, A. moschatus subspecies tuberosus and A. manihot. A set of 40 SSR primers were tested, of which 30 primers gave reproducible amplification which were used further for diversity analysis. With a mean of 7.1 bands per SSR, DNA amplification with 30 SSRs generated a total 213 bands, of which 60.66 % were recorded polymorphic. Polymorphic information content ranged between 0.11 and 0.80 with an average of 0.52, indicating that the majority of primers were informative. The Jaccard’s coefficient ranged from 0.107 to 0.969. The UPGMA analysis grouped Abelmoschus genotypes into three main clusters at a cut-off of 0.20. Results of present study revealed that sufficient variation exists among the studied accessions and GAO-5 which was found highly diverse can be exploited for okra improvement. The outcome of present research would assist to make use of Ablemoschus germplasm for okra breeding.     

Is It Still Necessary to Continue to Collect Crop Genetic Resources in the Mediterranean Area? A Case Study in Catalonia

Crop genetic resources have been extensively collected in Europe in the last century, creating large, publicly available ex situ collections. While this huge genetic diversity is often underutilized, in recent decades, several initiatives have emerged at the local level to collect germplasm cultivated on farm. Uncoordinated actors often carry out these collecting missions without considering previously collected data. To explore whether new collecting missions are likely to be worthwhile, we studied the crop genetic resources conservation network in Catalonia by analyzing the passport data and geographical distribution of germplasm stored in seed banks. Moreover, to determine whether this germplasm was representative of the diversity cultivated on farm, we performed new collecting missions in four randomly selected areas in the European Union’s Natura 2000 network and compared the results with the ex situ databases. Seed banks currently hold a large germplasm collection (2931 accessions), although most materials are conserved in private collections without regulated systems for seed regeneration and are not present as duplicates in the National Inventory. One important shortcoming of the ex situ network is that the germplasm conserved ex situ shows a low geographical coverage, representing only 35.3% of the municipalities in Catalonia. Our new missions allowed us to collect 234 accessions, mostly tomatoes (17.5%) and beans (16.2%). The ecological indicators’ richness (both at species (S) and variety (V) levels), total abundance (A), and the Shannon-Weaver diversity index calculated at species (H2, considering the different accessions of each variety as a single population) and variety levels (H3, considering the intra-varietal genetic diversity) were higher in the newly collected germplasm than in the ex situ collections, suggesting that seed banks do not accurately represent the genetic diversity still cultivated on farm. Moreover, some important landraces from each area were absent or underrepresented in the ex situ collections. Thus, it is necessary to continue to devote efforts to collecting germplasm; better organization between actors and targeting specific species/varieties can increase the efficiency of new collecting missions. As a conclusion, we propose different criteria to guide new missions and to improve the network’s conservation activities.     

Genetic diversity and population structure of trifoliate yam (<Emphasis Type="Italic">Dioscorea dumetorum</Emphasis> Kunth) in Cameroon revealed by genotyping-by-sequencing (GBS)

Background

Yams (Dioscorea spp.) are economically important food for millions of people in the humid and sub-humid tropics. Dioscorea dumetorum (Kunth) is the most nutritious among the eight-yam species, commonly grown and consumed in West and Central Africa. Despite these qualities, the storage ability of D. dumetorum is restricted by severe postharvest hardening of the tubers that can be addressed through concerted breeding efforts. The first step of any breeding program is bound to the study of genetic diversity. In this study, we used the Genotyping-By-Sequencing of Single Nucleotide Polymorphism (GBS-SNP) to investigate the genetic diversity and population structure of 44 accessions of D. dumetorum in Cameroon. Ploidy was inferred using flow cytometry and gbs2ploidy.

Results

We obtained on average 6371 loci having at least information for 75% accessions. Based on 6457 unlinked SNPs, our results demonstrate that D. dumetorum is structured into four populations. We clearly identified, a western/north-western, a western, and south-western populations, suggesting that altitude and farmers-consumers preference are the decisive factors for differential adaptation and separation of these populations. Bayesian and neighbor-joining clustering detected the highest genetic variability in D. dumetorum accessions from the south-western region. This variation is likely due to larger breeding efforts in the region as shown by gene flow between D. dumetorum accessions from the south-western region inferred by maximum likelihood. Ploidy analysis revealed diploid and triploid levels in D. dumetorum accessions with mostly diploid accessions (77%). Male and female accessions were mostly triploid (75%) and diploid (69%), respectively. The 1C genome size values of D. dumetorum accessions were on average 0.333?±?0.009?pg and 0.519?±?0.004?pg for diploids and triploids, respectively.

Conclusions

Germplasm characterization, population structure and ploidy are an essential basic information in a breeding program as well as for conservation of intraspecific diversity. Thus, results obtained in this study provide valuable information for the improvement and conservation of D. dumetorum. Moreover, GBS appears as an efficient powerful tool to detect intraspecific variation.

     

Genetic diversity and structure analyses on the natural populations of diploids and triploids of tiger lily, <Emphasis Type="Italic">Lilium lancifolium</Emphasis> Thunb., from Korea,China, and Japan

Tiger lily, Lilium lancifolium Thumb. is an endemic species in Korea. Genetic structure and diversity of diploid and triploid tiger lily accessions were analyzed using SSRP and IRAP molecular markers. All genetic indices analyzed showed that the diploids hold higher genetic variations compared to the triploids which were collected from Korea, China, and Japan. All variations found in triploids were also found in diploids to support the autotriploidy of L. lancifolium. It was also revealed that the triploid populations have experienced population regrowth from small population after population bottleneck. The population and phylogenetic analyses showed that analyzed tiger lilies were partitioned into three populations, Pop 1 from West Sea of Korean Peninsula, Pop 2 from South Sea of Korean Peninsula and Pop 3 of triploids in which many accessions of West Sea and South Sea were admixed in each partitioned group. Although the level of population differentiation is not extensive, F_ST analysis revealed some level of differentiation among the three populations. The analyses of genetic structure and phylogenetic dendrogram revealed admixture in some accessions among populations. All the results obtained in our analyses imply that autotriploid tiger lilies were originated from a small number of accessions of diploid L. lancifolium.     

Genetic diversity and population structure assessment of Chinese <Emphasis Type="Italic">Senna obtusifolia</Emphasis> L. by molecular markers and morphological traits of seed

Senna obtusifolia L. is an important medicinal plant in Asia. This study was the first report on the genetic diversity and population structure of S. obtusifolia which were collected from 47 geographic populations widespread in China. Inter-Simple Sequence Repeat (ISSR) and Start Codon Target Polymorphism (SCoT) combined with seeds morphological traits were used to investigate the relationship of 47 populations. 11 ISSR primers yielded 98 polymorphic bands with 81.67% polymorphism. 24 SCoT primers yielded 267 polymorphic bands with 89.59% polymorphism. The number of allele (Na), the number of effective allele (Ne), Nei’s diversity index (H), and Shannon’s information index (I) reflected a high level of genetic diversity of S. obtusifolia species. The greatest genetic distance (G _D) existed between Southwest and Northwest (0.4022_ISSR/0.5019_SCoT), while the Eastern and Northern showed the least genetic distance (0.1751_ISSR/0.2186_SCoT). The genetic differentiation (Gst) was 0.4875_ISSR/0.4434_SCoT, and the gene flow (Nm) was 0.5256_ISSR/0.6275_SCoT, which indicated that gene exchange among four regions was limited. 47 samples were divided into four clusters mainly according to their geographic distribution through clustering and structure analysis. The analysis on the combined data of ISSR and SCoT showed more reliable and superior results than single analysis of ISSR and SCoT. This study explored the effectiveness of ISSR and SCoT markers to evaluate the genetic diversity and population structure of S. obtusifolia and provided useful information for S. obtusifolia germplasm research and breeding program.