Genetic diversity assessment and ex situ conservation strategy of the endangered Dendrobium officinale (Orchidaceae) using new trinucleotide microsatellite markers

Dendrobium officinale (Orchidaceae) is an endangered plant species with important medicinal value. To evaluate the effectiveness of ex situ collection of D. officinale genetic diversity, we developed 15 polymorphic trinucleotide microsatellite loci of D. officinale to examine the genetic diversity and structure of three D. officinale germplasm collections comprising 120 individuals from its germplasm collection base and their respective wild populations consisting of 62 individuals from three provinces in China. The three germplasm collections showed reductions in gene diversity and average number of alleles per locus, but an increase in average number of rare alleles (frequency?≤?0.05) per locus in comparison to their wild populations. However, the differences in gene diversity between the germplasm collections and wild populations were not statistically significant. The analysis using STRUCTURE revealed evident differences in genetic composition between each germplasm collection and its wild population, probably because the D. officinale individuals with distinct genotypes in each wild population were unevenly selected for establishing its germplasm collection. For conservation management plans, we propose that D. officinale individuals with rare alleles need to be conserved with top priority, and those individuals with the most common alleles also should be concerned. The 15 new microsatellite loci may be used as a powerful tool for further evaluation and conservation of the genetic diversity of D. officinale germplasm resources.     

Association mapping of agro-morphological characters among the global collection of finger millet genotypes using genomic SSR markers

Identification of alleles responsible for various agro-morphological characters is a major concern to further improve the finger millet germplasm. Forty-six genomic SSRs were used for genetic analysis and population structure analysis of a global collection of 190 finger millet genotypes and fifteen agro-morphological characters were evaluated. The overall results showed that Asian genotypes were smaller in height, smaller flag leaf length, less basal tiller number, early flowering and early maturity nature, small ear head length, and smaller in length of longest finger. The 46 SSRs yielded 90 scorable alleles and the polymorphism information content values varied from 0.292 to 0.703 at an average of 0.442. The gene diversity was in the range of 0.355 to 0.750 with an average value of 0.528. The 46 genomic SSR loci grouped the 190 finger millet genotypes into two major clusters based on their geographical origin by the both phylogenetic clustering and population structure analysis by STRUCTURE software. Association mapping of QTLs for 15 agro-morphological characters with 46 genomic SSRs resulted in identification of five markers were linked to QTLs of four traits at a significant threshold (P) level of ≤0.01 and ≤0.001. The QTL for basal tiller number was strongly associated with the locus UGEP81 at a P value of 0.001 by explaining the phenotypic variance (R ²) of 10.8 %. The QTL for days to 50 % flowering was linked by two SSR loci UGEP77 and UGEP90, explained 10 and 8.7 % of R ² respectively at a P value of 0.01. The SSR marker, FM9 found to have strong association to two agro-morphological traits, flag leaf width (P—0.001, R ²—14.1 %) and plant height (P—0.001, R ²—11.2 %). The markers linked to the QTLs for above agro-morphological characters found in the present study can be further used for cloning of the full length gene, fine mapping and their further use in the marker assisted breeding programmes for introgression of alleles into locally well adapted germplasm.     

Population genetic structure of the tongue sole (Cynoglossus semilaevis) in Korea based on multiplex PCR assays with 12 polymorphic microsatellite markers

The tongue sole, Cynoglossus semilaevis, is an important fishery resource in Korea. About 100 tongue sole sampled from three major habitats along the western coast of Korea were assessed using multiplex assays with 12 highly polymorphic microsatellite loci to explore the population genetic structure of the species; 151 alleles and similar high levels of gene diversity (mean number of alleles (N_A) = 10.42, mean expected heterozygosity (He) = 0.78) were detected. Three populations showed significant Hardy–Weinberg equilibrium deviations at four loci. Although a significant difference in the number of unique alleles was observed among populations, genetic population subdivision was low by F-statistics (overall F _ST = 0.007, p < 0.05). However, this substructure was not supported by analysis of molecular variance or analyses of isolation by distance. The results suggest a lack of genetic structure among the tongue sole populations in Korean waters and that the populations should be managed as a single unit. The lack of genetic differentiation among samples may be due to high levels of larval dispersal in ocean currents. Alternatively, the populations may have diverged too recently for significant genetic differentiation to have become evident. Given the intensity of tongue sole aquaculture activity in China, which adjoins the western coast of Korea, the possibility that aquaculture may have partially contributed to the population genetic characteristics detected cannot be excluded. This study provides the basic information on nature population structure of C. semilaevis that may help to preserve and manage tongue soles in Korea.     

Identification of unique alleles and assessment of genetic diversity of rabi sorghum accessions using simple sequence repeat markers

Genetic diversity among 42 sorghum accessions representing landraces (19), advanced breeding lines (16), local cultivars (2) and release varieties (5) with 30 simple sequence repeat (SSR) markers revealed 7.6 mean number of alleles per locus showing 93.3% polymorphism and an average polymorphism information content of 0.78 which range from 0.22 (Xtxp12) and 0.91(Xtxp321). The average heterozygosity and effective number of alleles per locus were 0.8 and 6.65 respectively. Cluster analysis based on microsatellite allelic diversity clearly demarcated the accessions into ten clusters. A total of 24 unique alleles were obtained from seven SSR loci in 23 accessions in a size range of 110–380 bp; these unique alleles may serve as diagnostic tools for particular region of the genome of respective genotypes. Selected SSR markers from different linkage groups provided an accurate way of determining genetic diversity at the molecular level.     

Population genetic structure of the sea bass (Lateolabrax japonicus) in Korea based on multiplex PCR assays with 12 polymorphic microsatellite markers

Population genetics has been recognized as a key component of policy development for fisheries and conservation management. In this study, natural sea bass (Lateolabrax japonicus) populations in three ocean basins in Korea were assessed using multiplex assays with 12 highly polymorphic microsatellite loci; 203 alleles and similarly high levels of genetic diversity [mean number of alleles (N_A) = 14.43, mean expected heterozygosity (He) = 0.84] were detected. All populations showed significant heterozygote deficiency at four loci, which could be explained by the presence of null alleles. The genetic population subdivision was low and was significantly different according to F-statistics (overall F _ST = 0.003, R _ST = 0.005). However, this substructure was not supported by an analysis of molecular variance test, analyses of isolation by distance or Bayesian analysis. The passive dispersal of eggs/larvae via the main currents appears to facilitate gene flow. The possibility of a recent genetic bottleneck was observed in all three populations of L. japonicus, indicating that overfishing and degradation of the environment in recent years has led to a decline in the sea bass populations in Korea. Our study demonstrates that sea bass in Korea do not appear to be genetically partitioned and should be managed as a single unit; however, the potential for a rapid loss of genetic diversity remains. Information regarding the genetic characteristics of Korean sea bass populations has important implications for fishery management and conservation efforts and will aid in the sustainable exploitation of fishing resources and the preservation of biodiversity.     

Comparative Genetic Diversity of Wild and Captive Populations of the Bare-Faced Curassow (Crax fasciolata) Based on Cross-Species Microsatellite Markers: Implications for Conservation and Management

The bare-faced curassow (Crax fasciolata) is a large Neotropical bird that suffers anthropogenic pressure across much of its range. A captive population is maintained for conservation management, although there has been no genetic screening of stocks. Based on the six microsatellite markers developed for Crax globulosa, the genetic variability of C. fasciolata and possible differences between a wild and a captive population were investigated. Only three loci were polymorphic, with a total of 27 alleles. More than half of these alleles were private to the wild (n = 8) or captive (n = 7) populations. Significant deviations from Hardy–Weinberg equilibrium were restricted to the captive population. Despite the number of private alleles, genetic drift has probably promoted differentiation between populations. Our results indicate that wild C. fasciolata populations are genetically impoverished and structured, but species-specific microsatellite markers will be necessary for a more reliable assessment of the species’ genetic diversity.     

Genetic diversity in peach [Prunus persica (L.) Batsch] at the University of Florida: past,present and future

The University of Florida (UF) stone fruit breeding and genetics program was created in 1952 to develop early ripening stone fruit cultivars with high quality, adaptation to summer rainfall, low chilling requirements, and the ability to withstand high disease pressure. Diverse germplasm sources were used to introduce desirable traits in UF breeding pool. The main objective of this research was to determine the genetic diversity and population structure of the breeding germplasm, and to search for loci under selection. A total of 195 peach genotypes were used: UF cultivars and advanced selections (n?=?168), cultivars and selections from the UF-UGA-USDA joint breeding effort (n?=?13), landrace cultivars (n?=?4), high-chilling cultivars released by NCSU (n?=?5), and related Prunus (n?=?5) species. A total of 36 SSR markers distributed across the peach genome amplified 423 alleles. An average of 18 genotypes were detected per marker: A (number of observed alleles) of 11.43, Ae (effective number of alleles) of 2.58, Ho (observed heterozygosity) of 0.4, He (expected heterozygosity) of 0.52, F (Wright’s fixation index) of 0.25, and PIC (polymorphism information content) of 0.48. UPGMA cluster analysis based on Nei’s genetic distance represented best the known pedigree information for the germplasm pools. Two major groups were observed across the germplasm corresponding to melting and non-melting flesh cultivars/selections. Population structure results supported these two major groups. Several loci closely located to genome regions where different phenotypic traits have been previously mapped were detected to be under selection.     

Evaluation of SSR Markers for the Assessment of Genetic Diversity and Fingerprinting of Gossypium hirsutum Accessions

A total 177 simple sequence repeat (SSR) markers were screened using a set of 47 Upland cotton genotypes comprising 14 commercial varieties, 14 germplasm accessions and 19 advanced breeding lines to identify informative markers for genetic diversity assessment and fingerprinting in G. hirsutum. Only 21% (381177) of SSR markers tested showed polymorphism with a mean of 2.18 alleles per locus and with average polymorphism information content (PIC) of 0.32. The SSR markers revealed a Jaccard’ similarity coefficient ranging between 0.43 and 0.89, with an average of 0.67 among accessions. Cluster analysis using unweighted pair group method with arithmetic averages (UPGMA) and principal component analysis (PCA) indicated that majority of the genotypes were very closely related. All the 47 genotypes showed heterorygosity for at least one of the SSR loci. We discovered 19 rare and 6 unique alleles among the tested genotypes of cotton. Fingerprint based on all the 38 loci revealed a probability of identical match by chance of 3.98x10. A set of ten SSR markers was identified which could distinguish all the 47 genotypes with a moderate probability of identical match by chance (X?_D ⁿ = 0.01).     

De novo development and characterization of polymorphic microsatellite markers in a schilbid catfish, <Emphasis Type="Italic">Silonia silondia</Emphasis> (Hamilton, 1822) and their validation for population genetic studies

The stock characterization of wild populations of Silonia silondia is important for its scientific management. At present, the information on genetic parameters of S. silondia is very limited. The species-specific microsatellite markers were developed in current study. The validated markers were used to genotype individuals from four distant rivers. To develop de novo microsatellite loci, an enriched genomic library was constructed for S. silondia using affinity–capture approach. The markers were validated for utility in population genetics. A total number of 76 individuals from four natural riverine populations were used to generate data for population analysis. The screening of isolated repeat sequences yielded eleven novel polymorphic microsatellite loci. The microsatellite loci exhibited high level of polymorphism, with 6–24 alleles per locus and the PIC value ranged from 0.604 to 0.927. The observed (Ho) and expected (He) heterozygosities ranged from 0.081 to 0.84 and 0.66 to 0.938, respectively. The AMOVA analysis indicated significant genetic differentiation among riverine populations (overall F_ST = 0.075; P < 0.0001) with maximum variation (92.5 %) within populations. Cross-priming assessment revealed successful amplification (35–38 %) of heterologous loci in four related species viz. Clupisoma garua, C. taakree, Ailia coila and Eutropiichthys vacha. The results demonstrated that these de novo polymorphic microsatellite loci are promising for population genetic variation and diversity studies in S. silondia. Cross-priming results indicated that these primers can help to get polymorphic microsatellite loci in the related catfish species of family Schilbidae.     

Population structure and genetic diversity analysis of Indian and exotic rice (Oryza sativa L.) accessions using SSR markers

In order to understand the population structure and genetic diversity among a set of 82 rice genotypes collected from different parts of the Asian countries including India were characterized using 39 microsatellite loci. The Population structure analysis suggested that the optimum number of subpopulations was four (K = 4) among the rice genotypes, whereas phylogenetic analysis grouped them into three populations. The results obtained from phylogenetic and STRUCTURE analysis proved to be very powerful for the differentiation of rice genotypes based on their place of origin. The genetic diversity analysis using 39 SSR loci yielded 183 scorable alleles, out of which 182 alleles were observed to be polymorphic with an average of 4.8 alleles per locus. The Polymorphism Information Content (PIC) values for all the polymorphic primers across 82 rice genotypes varied from 0.02 to 0.77, with an average of 0.50. Gene diversity (He) was found to be in the range of 0.02 (RM484) to 0.80 (OSR13) with an average value of 0.55, while heterozygosity (Ho) was observed with an average of 0.07, ranging from 0.01 (RM334) to 0.31 (RM316). The present study resulted in identification of seven highly polymorphic SSR loci viz., OSR13, RM152, RM144, RM536, RM489, RM259 and RM271 based on the parameters like PIC value (≥0.70), gene diversity (≥0.71), and polymorphic alleles (≥6). These seven polymorphic primers can effectively be used in further molecular breeding programs and QTL mapping studies of rice since they exhibited very high polymorphism over other loci. SSR analysis resulted in a more definitive separation of clustering of genotypes indicating a higher level of efficiency of SSR markers for the accurate determination of relationships between accessions.     

Molecular characterization of Platonia insignis Mart. (“Bacurizeiro”) using inter simple sequence repeat (ISSR) markers

Platonia insignis Mart. (Clusiaceae) is widespread throughout the Amazon and adjacent areas. The fruits (known locally as “bacuri”) have significant commercial potential, but the species is under threat from agro-industrial expansion. The genetic variability within 72 genotypes of P. insignis belonging to ten populations collected in the Brazilian states of Maranhão and Piauí, and maintained in the germplasm collection of Embrapa Meio-Norte, has been determined, and the organization of genetic diversity within populations, between populations and among geographic groups verified. Eighteen selected inter simple sequence repeat primers allowed amplification of 236 loci of which 221 (93.64 %) were polymorphic, indicating a high level of genetic diversity. At the population level, the Shannon and Nei diversity indices ranged from 0.082 to 0.323 and from 0.120 to 0.480, respectively. The global coefficient of genetic differentiation (G _ST ) was 0.4730 indicating that differentiation between populations was significant, a finding that was confirmed by analysis of molecular variance (Φ _ST  = 0.28). UPGMA cluster analysis revealed that the genotypes could be stratified into groups that were well defined and consistent with those identified in the dendrogram constructed using pair wise Φ _ST values. The high genetic diversity established in this study may facilitate the management and conservation of the germplasm of P. insignis.     

Genetic diversity,population structure and association analysis in linseed (<Emphasis Type="Italic">Linum usitatissimum</Emphasis> L.)

The present investigation aimed to explore the level of genetic diversity, determine the population structure in a larger set of germplasm of linseed using microsatellite marker and identify linked markers through association mapping. A total of 168 accessions of linseed were evaluated for major agro-economic traits and SSRs markers deployed for diversity assessment. A total of 337 alleles were amplified by 50 SSRs ranging from 2 to 13 with an average of 6.74 ± 2.8 alleles per loci. The neighbor joining based clustering grouped all the accessions into three major clusters that were also confirmed by scatter plot of PCoA. While model based clustering determined four sub-populations (K = 4). Further, analysis of molecular variance analysis considering three population showed that maximum variation (79%) was within the population. We identified one putative SSR marker (Lu_3043) linked with days to 50% flowering through both GLM and MLM analysis of association mapping. The results of this preliminary study revealed genetic diversity, population structure in linseed and linked marker which could be utilized in future breeding program.     

Exploring genetic variability within lentil (Lens culinaris Medik.) and across related legumes using a newly developed set of microsatellite markers

Lentil (Lens culinaris Medik.) is an economically important grain legume, yet the genetic and genomic resources remain largely uncharacterized and unexploited in this crop. Microsatellites have become markers of choice for crop improvement applications. Hence, simple sequence repeat (SSR) markers were developed for lentil through the construction of genomic library enriched for GA/CT motifs. As a result 122 functional SSR primer pairs were developed from 151 microsatellite loci and validated in L. culinaris cv. Precoz. Thirty three SSR markers were utilized for the analysis of genetic relationships between cultivated and wild species of Lens and related legumes. A total of 123 alleles were amplified at 33 loci ranging from 2–5 alleles with an average of 3.73 alleles per locus. Polymorphic information content (PIC) for all the loci ranged from 0.13 to 0.99 with an average of 0.66 per locus. Varied levels of cross genera transferability were obtained ranging from 69.70 % across Pisum sativum to 12.12 % across Vigna radiata. The UPGMA based dendrogram was able to establish the uniqueness of each genotype and grouped them into two major clusters clearly resolving the genetic relationships within lentil and related species. The new set of SSR markers reported here were efficient and highly polymorphic and would add to the existing repertoire of lentil SSR markers to be utilized in molecular breeding. Moreover, the improved knowledge about intra- and inter-specific genetic relationships would facilitate germplasm utilization for lentil improvement.     

Genetic analysis of forest species <Emphasis Type="Italic">Eugenia uniflora</Emphasis> L. through of newly developed SSR markers

Nine microsatellite loci for genetic analysis of three populations of the tropical tree Eugenia uniflora L. (pitanga or Brazilian cherry) from fragments of semideciduous forest were developed. We used the technique of building a (GA) _n and (CA) _n microsatellite-enriched library by capture with streptavidin-coated magnetic beads. We assessed the polymorphism of seven microsatellites in 84 mature trees found in three areas (Ribeirão Preto, Tambaú and São José do Rio Pardo), highly impacted by the agricultural practices, in a large region among Pardo river and Mogi-Guaçu river basins, in state of São Paulo, Brazil. All loci were polymorphic, and the number of alleles was high, ranging from 6 to 24, with a mean of 14.4. All stands showed the same high level of genetic diversity (mean H _E  = 0.83) and a low genetic differentiation (mean F _ST = 0.031), indicating that genetic diversity was higher within rather than among populations. Seven of the nine loci were highly variable, and sufficiently informative for E. uniflora. It was concluded that these new SSR markers can be efficiently used for gene flow studies.     

SSR-based population structure, molecular diversity and linkage disequilibrium analysis of a collection of flax (Linum usitatissimum L.) varying for mucilage seed-coat content

Flax seed mucilage (SM) presents specific biological activities useful for the food and pharmaceutical industries. Understanding the population structure, genetic diversity and linkage disequilibrium (LD) of germplasm varying for mucilage content is pivotal for the identification of genes and quantitative trait loci underlying mucilage variation by association mapping (AM). In this study, 150 microsatellite loci were used to assess the population structure, genetic diversity and LD of a set of 60 flax cultivars/accessions capturing the breadth of SM variation in flax germplasm. STRUCTURE analysis and similarity-based methods revealed the presence of three populations reflecting mainly their geographic origins (South Asia, South America and North America), and the impact of germplasm exchange within and between North American flax breeding programs. Analysis of molecular variance showed that 78.32% of the genetic variation resided within populations and 21.68% among populations. The phi-statistic (??_st) value of 0.22 confirmed the presence of a strong population structure. A total of 408 alleles were detected, with the South American population capturing the highest overall diversity. However, the genetic diversity was narrow, as indicated by the small number of alleles per locus (2.72) and gene diversity (mean?=?0.34). LD was significant between 3.9% (r ²) and 36.2% (D??) of the loci pairs (FDR?<?0.05). The mean r ² and D?? were 0.26 and 0.53, respectively. The results suggest that the collection could be useful in AM studies aimed at the discovery of genes/alleles involved in SM; however a greater diversity may be required to improve the AM resolution.     

Characterization of polymorphic microsatellite markers and genetic diversity in wild bronze featherback, Notopterus notopterus (Pallas, 1769)

Six polymorphic microsatellite DNA loci were identified in the primitive fish, bronze featherback, Notopterus notopterus for the first time and demonstrated significant population genetic structure. Out of the six primers, one primer (NN90) was specific to N. notopterus (microsatellite sequence within the RAG1 gene) and five primers were product of successful cross-species amplification. Sixty-four primers available from 3 fish species of order Osteoglossiformes and families Notopteridae and Osteoglossidae were tested to amplify homologous microsatellite loci in N. notopterus. Fifteen primer pairs exhibited successful cross-priming PCR product. However, polymorphism was detected only at five loci. To assess the significance of these six loci (including NN90) in population genetic study, 215 samples of N. notopterus from five rivers, viz Satluj, Gomti, Yamuna, Brahmaputra and Mahanadi were analyzed. The five sample sets displayed different diversity levels and observed heterozygosity ranged from 0.6036 to 0.7373. Significant genotype heterogeneity (P < 0.0001) and high F_ST (0.2205) over all loci indicated that the samples are not drawn from the same genepool. The identified microsatellite loci are promising for use in fine-scale population structure analysis of N. notopterus.     

Development and characterization of genomic SSR markers in Cynodon transvaalensis Burtt-Davy

Simple sequence repeat (SSR) markers are a major molecular tool for genetic and genomic research that have been extensively developed and used in major crops. However, few are available in African bermudagrass (Cynodon transvaalensis Burtt-Davy), an economically important warm-season turfgrass species. African bermudagrass is mainly used for hybridizations with common bermudagrass [C. dactylon var. dactylon (L.) Pers.] in the development of superior interspecific hybrid turfgrass cultivars. Accordingly, the major objective of this study was to develop and characterize a large set of SSR markers. Genomic DNA of C. transvaalensis ‘4200TN 24-2’ from an Oklahoma State University (OSU) turf nursery was extracted for construction of four SSR genomic libraries enriched with [CA] _n , [GA] _n , [AAG] _n , and [AAT] _n as core repeat motifs. A total of 3,064 clones were sequenced at the OSU core facility. The sequences were categorized into singletons and contiguous sequences to exclude redundancy. From the two sequence categories, 1,795 SSR loci were identified. After excluding duplicate SSRs by comparison with previously developed SSR markers using a nucleotide basic local alignment tool, 1,426 unique primer pairs (PPs) were designed. Out of the 1,426 designed PPs, 981 (68.8 %) amplified alleles of the expected size in the donor DNA. Polymorphisms of the SSR PPs tested in eight C. transvaalensis plants were 93 % polymorphic with 544 markers effective in all genotypes. Inheritance of the SSRs was examined in six F₁ progeny of African parents ‘T577’ × ‘Uganda’, indicating 917 markers amplified heritable alleles. The SSR markers developed in the study are the first large set of co-dominant markers in African bermudagrass and should be highly valuable for molecular and traditional breeding research.     

Genetic diversity and structure of an endemic and critically endangered stream river salamander (Caudata: Ambystoma leorae) in Mexico

Small or isolated populations are highly susceptible to stochastic events. They are prone and vulnerable to random demographic or environmental fluctuations that could lead to extinction due to the loss of alleles through genetic drift and increased inbreeding. We studied Ambystoma leorae an endemic and critically threatened species. We analyzed the genetic diversity and structure, effective population size, presence of bottlenecks and inbreeding coefficient of 96 individuals based on nine microsatellite loci. We found high levels of genetic diversity expressed as heterozygosity (H_o = 0.804, H_e = 0.613, H_e* = 0.626 and H_Nei = 0.622). The population presents few alleles (4–9 per locus) and genotypes (3–14 per locus) compared with other mole salamanders species. We identified three genetically differentiated subpopulations with a significant level of genetic structure (F_ST = 0.021, R_ST = 0.044 y D_est = 0.010, 95 % CI). We also detected a reduction signal in population size and evidence of a genetic bottleneck (M = 0.367). The effective population size is small (Ne = 45.2), but similar to another mole salamanders with restricted distributions or with recently fragmented habitat. The inbreeding coefficient levels detected are low (F_IS = ?0.619–0.102) as is gene flow. Despite, high levels of genetic diversity A. leorae is critically endangered because it is a small isolated population.     

Population genetics of the Federally Threatened Miccosukee gooseberry (Ribes echinellum), an endemic North American species

Ribes echinellum (Coville) Rehder (Miccosukee gooseberry; Grossulariaceae) is a Federally Threatened species known from only two localities: Jefferson County (Florida, FL) and McCormick County (South Carolina, SC). This perennial shrub, ca. 1 m tall, is deciduous, and reproduces both vegetatively (clonal growth) and sexually (seed production). Recent surveys of the FL population revealed a dramatic decline in plant numbers. To assist in conservation and management of this species in FL and SC populations, microsatellite genetic markers were used to identify genotypes and assess the genetic structure of R. echinellum. We genotyped seven microsatellite loci in 102 individuals: 74 collected in FL and 28 in SC. Unbiased heterozygosity was between 0.28 and 0.53. All seven loci were polymorphic, showing a range of 1.52–2.13 effective number of alleles per locus (mean = 1.75). The two populations of R. echinellum show low genetic diversity, especially in SC. Clonality was not widespread, but was higher in the SC population. Both populations show signatures of bottlenecks but isolation by distance was not evident. We found significant deviation from HW equilibrium, with higher number of heterozygotes than expected. However when HW test was done for the combined populations as two separate groups, only FL showed a significant HW test and for SC the test was non-significant. Bayesian analysis and F_ST values suggest high genetic divergence between the populations. These results are important for developing a recovery plan and an ex situ and reintroduction conservation programs.