Genetic Diversity and Phylogenetic Relationships of Two Closely Related Northeast China <Emphasis Type="Italic">Vicia</Emphasis> Species Revealed with RAPD and ISSR Markers

RAPD and ISSR analyses revealed genetic diversity and relationships among 11 populations of two closely related northeast China Vicia species, Vicia ramuliflora and V. unijuga. Both methods yielded similar and complementary results, showing high genetic diversity. Vicia ramuliflora had 100% polymorphic loci in both RAPD and ISSR, and V. unijuga had 100% polymorphic loci for RAPD and 98.96% for ISSR. Genetic differentiation was moderate among populations of each species. Genetic variation was distributed mainly within populations for the two species. The high level of gene flow was important for the allocation of genetic variation. The UPGMA dendrogram and principal coordinates analysis at the level of individuals and populations showed that V. ramuliflora and V. unijuga were more closely related than either of them was to the outgroup species, V. cracca. The small molecular variance of V. ramuliflora and V. unijuga supports the conclusion that these two species had a common ancestor.     

Genetic structure of the Russian populations of <Emphasis Type="Italic">Pyrenophora tritici-repentis</Emphasis>, determined by using microsatellite markers

The population genetic structure of plant pathogenic fungus Pyrenophora tritici-repentis was examined using microsatellite (SSR) markers. According to the geographical origin of the pathogen populations, they were designated as North Caucasian (S, 33 isolates), northwest (Nw, 39), and Omsk (Om, 43). The populations were analyzed at the nine most polymorphic SSR loci, at which 75 alleles were identified. To characterize the genetic variation within and between populations, the AMOVA algorithm as implemented in the Arlequin v. 3.5 software program was used. The number of alleles per locus ranged from 5 to 12 and their sizes varied within the range from 180 to 400 bp. The mean gene diversity at SSR loci was high for all populations (H = 0.58–0.75). The populations were considerably different in the frequencies of individual alleles of the SSR loci. Most isolates in the populations were represented by unique haplotypes. The within-population variation of the isolates at molecular markers was 86.4%; among the populations, 13.6%. Substantial interpopulation differences were found between the Om and S (F_st = 0.16) and between the Om and Nw (F_st = 0.20) populations, whereas between the S and Nw populations, these differences were small (F_st = 0.05). Thus, it was demonstrated that the population of P. tritici-repentis from Omsk oblast had the independent status of the geographical population; northwest and North Caucasian populations differed in the allelic diversity of SSR loci, and despite the low F_st value (0.05), they also belonged to independent geographical populations.     

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.     

Genetic diversity of the African poplar (<Emphasis Type="Italic">Populus ilicifolia</Emphasis>) populations in Kenya

We evaluated the genetic diversity of the African poplar (Populus ilicifolia) populations found in Kenya compared with reference samples of five poplar species from North America and one species introduced in Kenya from India (KEFRI-Kenya). Amplified fragment length polymorphism (AFLP) was used with the objective of providing important information for breeding and in situ/ex situ conservation of this species. Samples collected from three locations along the species’ natural range (Athi, Ewaso Nyiro, and Tana rivers) were compared with four samples of locally planted Populus deltoides stand introduced from India and ten reference samples from North America. Six AFLP primer combinations produced 521 clear bands for analysis. The percentage polymorphic loci were lowest in Tana (20.4 %) and highest in Athi (40.6 %). The average heterozygosity across the studied populations was between 0.07 and 0.3. AMOVA revealed more genetic variation partitioning within population (87 %; P?<?0.01) than among populations (13 %; P?<?0.01) suggesting significant genetic variation between populations. Further, UPGMA delineation showed two clusters of the Tana, Athi, and Ewaso Nyiro populations clustered together compared to the North America and India/KEFRI reference samples. Moreover, the study showed that the Athi population is more diverse than those of Tana and Ewaso Nyiro and may be important for conservation, domestication, and improvement studies. The genetic differentiation (F _ST ?=?0.134) among Kenyan P. ilicifolia populations suggests limited possibility of gene flow between these populations.     

Isolation of Polymorphic RAPD-SSR Markers from Xinjiang Arctic Grayling (<Emphasis Type="Italic">Thymallus arcticus grubei</Emphasis>) and a Test of Cross-Species Amplification

A total of 18 polymorphic microsatellite loci were isolated and characterized from RAPD products in the Xinjiang Arctic Grayling (Thymallus arcticus grubei). The number of alleles (N_a) per locus varied from 2 to 10. Observed (H_o) and expected (H_e) heterozygosities ranged from 0.64 to 0.92, and from 0.63 to 0.88, respectively. Considerable differences were found among HBH, FH and FY populations in the number of alleles, effective number of alleles, number of genotypes at all of these loci. These new RAPD-SSR markers have provided a helpful tool for genetic analyses and resources conservation of T. arcticus grubei. Five additional fish species, Amur grayling (Thymallus grubii), Taimen (Hucho taimen), Sea perch (Lateolabrax japonicus), Lenok (Brachymystax lenok) and Red seam bream (Pagrosomus major) were assessed for cross-species amplification. Three of the five species showed at least one polymorphic locus. In addition, seven loci were found to be polymorphic in at least one species.     

In silico polymorphic novel SSR marker development and the first SSR-based genetic linkage map in pistachio

Simple sequence repeats (SSRs) are co-dominant markers, and are very useful in constructing consensus maps in heterozygous perennial plant species like pistachio. Pistacia vera L. is the only cultivated species in the genus Pistacia. It is dioecious with a haploid chromosome count of n =?15. Saturated genetic linkage maps can be a reference to identify markers linked to economically important phenotypic traits that could be useful for early breeding and selection programs. Therefore, this study aimed to develop polymorphic SSR markers in silico and to construct the first SSR-based genetic linkage map in pistachio. The DNA sequences of three cultivars (Siirt, Ohadi, and Bagyolu) of P. vera and one genotype belonging to P. atlantica (Pa-18) were obtained by next-generation sequencing, and 625 polymorphic SSR loci were identified from 750 screened in silico polymorphic SSR primer pairs. The novel SSRs were used to construct SSR-based genetic linkage maps in pistachio along with published SSRs in Siirt × Bagyolu F1 population. Most (71.4%) of the SSRs were common markers that were used to construct consensus and parental maps spanning 15 linkage groups (LGs). A total of 384, 317, and 341 markers were mapped in the consensus, female, and male genetic maps with total lengths of 1511.3, 1427.0, and 1453.4 cM, respectively. The large number of SSR markers discovered and the first SSR-based genetic linkage map constructed in this study will be useful for anchoring loci for map integration, and will facilitate marker-assisted selection efforts for important horticultural traits in the genus Pistacia.     

A high-density genetic linkage map of bronze loquat based on SSR and RAPD markers

We constructed a high-density genetic linkage map of bronze loquat (Eriobotrya deflexa) by using a three-way cross of loquat (Eriobotrya japonica) × (loquat × bronze loquat) and simple sequence repeat (SSR) and random amplified polymorphic DNA (RAPD) markers. The positions of the SSR loci used in this study were previously identified on reference maps of pears (Pyrus spp.) and apples (Malus spp.). The map of bronze loquat (‘Taiwan loquat No. 1’) consisted of 308 loci including 167 SSRs (8 loquat, 57 pear, and 102 apple SSRs), 140 RAPDs, and the loquat canker resistance gene Pse-a on 19 linkage groups covering a genetic distance of 1036 cM. Almost all loquat linkage groups were aligned to the pear consensus map by using at least two pear or apple SSRs, suggesting that positions and linkages of SSR loci were well conserved between loquat and pear and between loquat and apple. The constructed map may be used to determine the location of genes and quantitative trait loci of interest and to analyze genome synteny in the tribe Pyreae, subfamily Spiraeoideae of the family Rosaceae.     

Analysis of microsatellite loci variability in rare and endemic species <Emphasis Type="Italic">Allium regelianum</Emphasis> A.K. Becker ex Iljin

SSR analysis of rare and endemic species Allium regelianum, which grows in the south of Russia, was performed for the first time. Variability analysis of 88 accessions of A. regelianum was carried out using four highly polymorphic microsatellite loci (PIC value ranged from 0.55 to 0.72). SSR-analysis made it possible to revealed polymorphism within and among the populations of A. regelianum from Volgograd region. Analysis of Wright’s F-statistics and the analysis of molecular variance showed that more than 90% of total genetic variation of the species was attributed to the differences within the populations and less than 10% of the differences were found among the populations. Cluster analysis of 46 accessions of A. regelianum from three populations of Volgograd region and principal coordinate analysis of all accessions did not reveal differentiation among the populations.     

Variability of nuclear microsatellite loci in the populations of Siberian dwarf pine (<Emphasis Type="Italic">Pinus pumila</Emphasis> (Pallas) Regel) from the Russian part of the range

Variability of nuclear microsatellite loci was examined in Siberian dwarf pine. Six microsatellite loci (RPS2, RPS6, RPS12, RPS124, RPS127, Pc18) demonstrated different polymorphism levels in ten populations of Siberian dwarf pine. The average number of alleles per locus was 4.88, the average observed heterozygosity was 0.465, and the average expected heterozygosity was 0.510. About 13% of total genetic variability was explained by the genetic differences between the populations (F _ST = 0.129). Genetic distances between the examined populations of Pinus pumila inferred from the data on the SSR marker frequencies statistically significantly correlated with the geographical distances between the population samples. The level of genetic variability of the populations from Kamchatka Peninsula was lower than that demonstrated by continental and island populations. The genetic differentiation of the Kamchatka–Magadan and other populations of Siberian dwarf pine observed in our study can be explained in terms of their formation from different Pleistocene refugial centers.     

<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.     

Morphological and genetic differentiation among four pigment producing Indian species of <Emphasis Type="Italic">Phoma</Emphasis> (Saccardo, 1899)

A PCR-based technique, involving the random amplification of polymorphic DNA (RAPD), was used for assessing genetic relatedness among isolates of the genus Phoma. Randomly Amplified Polymorphic DNA (RAPD) revealed the presence of interspecific genetic variation among the pigment producing isolates of Phoma and has shown distinct phylogenetic cluster. The major objective of the study was to study the genetic variation, if any. Study was aimed to differentiate four pigment producing species of Phoma based on morphological studies and molecular markers in general and RAPD in particular. We found that the test species of Phoma can be very well differentiated using molecular markers. Phoma sorghina was differentiated from P. exigua, P. fimeti and P. herbarum. RAPD profiles of P. herbarum and P. fimeti has shown the maximum similarity, which indicates the genetic relatedness among these two species which were considered earlier as distinct species based on morphological observation.     

Polymorphism and differentiation of multilocus DNA markers in natural populations of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Using multilocus (RAPD) markers, variation and divergence of genomic DNA was examined in two Drosophila melanogaster populations from Russia and three populations from Ukraine. The populations were found to exhibit high polymorphism at RAPD markers. Estimation of genetic distances between the populations showed low differentiation of geographically distant populations of D. melanogaster. Significant gene flow between the D. melanogaster populations was found, which depended on the geographical distance between them.     

Genetic differentiation among natural populations of the lizard complex <Emphasis Type="Italic">Darevskia raddei</Emphasis> as inferred from genome microsatellite marking

The article presents the genetic parameters of the populations of lizards of the Darevskia raddei complex (D. raddei nairensis and D. raddei raddei) and the populations of D. valentini calculated on the basis of the analysis of variability of 50 allelic variants of the three nuclear genome microsatellite-containing loci of 83 individuals. It was demonstrated that the F_st genetic distances between the populations of D. raddei nairensis and D. raddei raddei were not statistically significantly different from the F_st genetic distances between the populations of different species, D. raddei and D. valentini. At the same time, these distances were statistically significantly higher than the F_st distances between the populations belonging to one species within the genus Darevskia. These data suggest deep divergence between the populations of D. raddei raddei and D. raddei nairensis of the D. raddei complex and there arises the question on considering them as separate species.     

Genetic structure of the populations of <Emphasis Type="Italic">Dactylorhiza ochroleuca</Emphasis> and <Emphasis Type="Italic">D. incarnata</Emphasis> (Orchidaceae) in the area of their joint growth in Russia and Belarus

We carried out an allozyme analysis to investigate polymorphism and genetic structure of the populations of D. incarnata and D. ochroleuca in regions of their joint growth in Russia and Belarus. We found that D. ochroleuca individuals in the populations of the Urals and Siberia, which are distant fragments from the main range of the species, do not differ significantly from individuals within the main part of the area (Belarus) on the basis of the allelic composition of eight gene loci. We revealed that D. ochroleuca and D. incarnata are differentiated by different alleles of the GDH locus. Thus, we established a genetic marker suitable to distinguish these closely related taxa. In addition to the GDH locus, D. ochroleuca and D. incarnata in the places of their joint growth, differ in the allelic structure of the PGI and NADHD loci. D. incarnata from the Urals and Siberia were polymorphic for both loci, and individuals from Belarus were polymorphic for one locus (PGI). In contrast, all D. ochroleuca individuals growing in sympatric populations with polymorphic D. incarnata were homozygous for the same alleles. Thus, comparison of the genetic structure of D. ochroleuca and D. incarnata points to the existence of a genetic isolation and a functioning isolation mechanism even under conditions of their joint growth. We found that the GDH locus in D. incarnata is polymorphic only in populations which grow together with D. ochroleuca, with exception a few examples. Thus, we conclude that variability of the GDH locus in D. incarnata is associated with hybridization with D. ochroleuca.     

Negligence in the Atlantic forest,northern Brazil: a case study of an endangered orchid

Currently, many Brazilian orchids are threatened with extinction resulting from habitat loss and intense harvesting pressure stemming from their value as ornamental plants. Therefore, the genetic diversity in remaining populations is fundamental to the survival of these species in natural environments. In order to inform conservation strategies, this study evaluated the genetic diversity and structure of Cattleya granulosa populations. The sample consisted of 151 individuals from 12 populations in the Atlantic Forest, northeastern Brazil, evaluated using 91 ISSR markers. Genetic variability was assessed through molecular variance, diversity indexes, clusters of genotypes through Bayesian analysis, and tests for genetic bottlenecks. From all polymorphic loci, genetic diversity (H_E) varied between 0.210 and 0.321 and the Shannon index ranged from 0.323 and 0.472. Significant genetic differentiation between populations (Φ_ST = 0.391; P < 0.0001) resulted in the division of the populations into five groups based on the log-likelihood Bayesian analysis. We found significant positive correlation between geographical and genetic distances between populations (r = 0.794; P = 0.017), indicating isolation by distance. Patterns of allelic diversity within populations suggest the occurrence of bottlenecks in most C. granulosa populations (n = 8). Therefore, in order to maintain the genetic diversity of the species, the conservation of spatially distant groups is necessary.     

Molecular mapping of the genomic region conferring resistance to soybean stem canker in Hutcheson soybean

Genetic resistance to soybean stem canker, caused by the fungus Diaporthe phaseolorum var. meridionalis (Dpm), is controlled by five major, dominant, nonallelic genes Rdm1 to Rdm5. A genomic region containing the Rdm4 and Rdm5 genes was first described in Hutcheson soybean, where they were found to confer specific resistance to Argentinean physiological races of Dpm. Here, we report the genetic mapping of Rdm4 and Rdm5 loci using two pheno- and genotypically characterized F_2:3 populations derived from Hutcheson cultivar. The mapping populations were screened with amplified fragment length polymorphism (AFLP) markers using bulk segregant analysis, and with simple sequence repeat (SSR) markers. Linkage analysis indicated that the Rdm4 and Rdm5 resistance loci were located in a genomic region collinear with the molecular linkage group (MLG) A2 (chromosome 8) of the soybean genetic map. The linkage group contains two SSR markers, Sat_162 and Satt233, flanking the Rdm4 and Rdm5 loci. These SSR will be useful to increase the efficiency of selection in breeding programs aimed to incorporate Rdm4 and Rdm5 genes into soybean elite germplasm.     

Population genetic diversity and geographical differentiation of MHC class II DAB genes in the vulnerable Chinese egret (<Emphasis Type="Italic">Egretta eulophotes</Emphasis>)

Major histocompatibility complex (MHC) genes are excellent markers for the study of adaptive genetic variation occurring over different geographical scales. The Chinese egret (Egretta eulophotes) is a vulnerable ardeid species with an estimated global population of 2600–3400 individuals. In this study, we sampled 172 individuals of this egret (approximately 6 % of the global population) from five natural populations that span the entire distribution range of this species in China. We examined their population genetic diversity and geographical differentiation at three MHC class II DAB genes by identifying eight exon 2 alleles at Egeu-DAB1, eight at Egeu-DAB2 and four at Egeu-DAB3. Allelic distributions at each of these three Egeu-DAB loci varied substantially within the five populations, while levels of genetic diversity varied slightly among the populations. Analysis of molecular variance showed low but significant genetic differentiation among five populations at all three Egeu-DAB loci (haplotype-based ?_ST: 0.029, 0.020 and 0.042; and distance-based ?_ST: 0.036, 0.027 and 0.043, respectively; all P < 0.01). The Mantel test suggested that this significant population genetic differentiation was likely due to an isolation-by-distance pattern of MHC evolution. However, the phylogenetic analyses and the Bayesian clustering analysis based on the three Egeu-DAB loci indicated that there was little geographical structuring of the genetic differentiation among five populations. These results provide fundamental population information for the conservation genetics of the vulnerable Chinese egret.     

Investigating the genetic diversity and differentiation patterns in the <Emphasis Type="Italic">Penstemon scariosus</Emphasis> species complex under different sample sizes using AFLPs and SSRs

Habitat fragmentation due to anthropogenic activities is the major cause of biodiversity loss. Endemic and narrowly distributed species are the most susceptible to habitat degradation. Penstemon scariosus is one of many species whose natural habitat is vulnerable to industrialization. All varieties of P. scariosus (P. scariosus var. albifluvis, P. scariosus var. cyanomontanus, P. scariosus var. garrettii, P. scariosus var. scariosus) have small distribution ranges, but only P. scariosus var. albifluvis is being considered for listing under the Endangered Species Act. We used eight microsatellites or simple sequence repeats (SSRs) loci and two amplified fragment length polymorphism (AFLP) primer combinations to investigate the population genetic structure and diversity of P. scariosus varieties. Moreover, we compared the utility of the two marker systems in conservation genetics and estimated an appropriate sample size in population genetic studies. Genetic differentiation among populations based on F_st ranged from low to moderate (F_st?=?0.056–0.157) and from moderate to high when estimated with D_es (D_es?=?0.15–0.32). Also, AMOVA analysis shows that most of the genetic variation is within populations. Inbreeding coefficients (F_is) were high in all varieties (0.20–0.56). The Bayesian analysis, STRUCTURE, identified three clusters from SSR data and four clusters from AFLPs. Clusters were not consistent between marker systems and did not represent the current taxonomy. MEMGENE revealed that a high proportion of the genetic variation is due to geographic distance (R²?=?0.38, P?=?0.001). Comparing the genetic measurements from AFLPs and SSRs, we found that AFLP results were more accurate than SSR results across sample size when populations were larger than 25 individuals. As sample size decreases, the estimates become less stable in both AFLP and SSR datasets. Finally, this study provides insight into the population genetic structure of these varieties, which could be used in conservation efforts.     

Analysis of genetic diversity among medicinal therapist <Emphasis Type="Italic">Trigonella foenum</Emphasis>-<Emphasis Type="Italic">graecum</Emphasis> L. genotypes through RAPD and SSR markers

Trigonella is recognized as a medicinal therapist throughout the globe due to its multifaceted rare medicinal properties. It is indigenous from Iran to Northern India but has gained global acceptance towards cultivation and consumption for its yellow-to-amber colored seed which substantially contributes to food, pharmaceutical, nutraceutical and cosmetic industry. Genetic diversity serves as an excellent tool for developing improved crop varieties with breeder preferred traits. Unfortunately, very little information available on variability existing in commercial Trigonella genotypes considerably impedes the crop improvement. In this study, ninety Trigonella genotypes belonging to most productive North Indian states were subjected to multilocus genotyping using RAPD (49) and SSR (13) primers and detected an average of 55.60 and 50.16% polymorphism, respectively. The percentage polymorphism range (RAPD, 16.7–90.90; SSR, 33.30–66.66) average band informativeness (RAPD, 0.182–0.85; SSR, 0.21–0.91) and resolving power (RAPD, 0.95–9.984; SSR, 1.68–7.28) obtained revealed the wide range of diversity prevailing among these genotypes. Hierarchical clustering of genotypes in nine different clusters showed Trigonella’s genetic variability has wide genetic distribution across different agro-climatic zones. No consistency was observed while grouping Trigonella varieties based on eco-geographical region. Eventually, knowledge of these genetic differences significantly contributes in designing intra-specific crosses with potential interest to spice breeding programs. To the best of our knowledge, this is the first report of genetic diversity using SSR molecular markers in Trigonella foenum-graecum L.     

Population study of the Udmurt population: Analysis of ten polymorphic DNA loci of the nuclear genome

Genetic structure of Southern and Northern ethnographic groups of the Udmurt population from six regions of the Republic of Udmurtia has been studied. All the samples were examined using ten polymorphic DNA loci: VNTR/PAH, VNTR/ApoB, VNTR/DATI, VNTR/eNOS, ACE, CCR5 Δ32, KM19, IVS6a, THOI, and FABP2. Allelic and genotype frequencies were estimated for each of the six populations. The average heterozygosity for these ten polymorphic loci varied from 0.47 in Udmurts from Glazovskii region to 0.53 in Udmurts from Malopurginskii region. The level of genetic variation (F _ST) between populations of Udmurts was 0.0048. Ethnographic subdivision of the population into Northern and Southern Udmurts is in good agreement with the values of genetic distances and phylogenetic analysis.