Population genetic variation in rare and endangered Iliamna (Malvaceae) in Virginia

Random amplified polymorphic DNA (RAPD) markers were used as input for an analysis of molecular variance (AMOVA), homogeneity of molecular variance analysis (HOMOVA), and cluster analysis to describe the population genetic structure of Iliamna corei, a federally endangered plant located only in Virginia, and I. remota , a rare plant in Virginia, Indiana, and Illinois. The analysis was performed to help clarify the taxonomic relationship between the two closely related species. We analysed four clones in the only known population of I. corei , breeding stock derived from seeds originating from the population site, and three I. remota populations in Virginia. Eighty-five percent of screened primers revealed DNA polymorphisms in Iliamna. Ninety-nine informative markers were generated using seven primers. No significant statistical differences (at P = 0.05) in RAPD variation was found between species (24% of variance) using the AMOVA procedure. However, within species/among populations (31 % of the variance) and within populations (45% of the variance) there were significant differences (P < 0.002). An unweighted paired group method using arithmetic averages (UPGMA) cluster analysis showed the federally endangered I. corei population to be genetically distinct from the apparently recently introduced (in Virginia: ∼ 100 ybp) I. remota. The lack of significant differences from the AMOVA and the high number shared bands between I. corei and I. remota suggest that I. corei may be more appropriately classified as a subspecies of I. remota. Iliamna corei plants in the natural population were genetically similar to one another while the I. corei breeding stock plants and I. remota plants were genetically relatively diverse.     

Patterns of genetic variation detected by RAPDs suggest a single origin with subsequent mutations and long-distance dispersal in the apomictic fern Dryopteris remota (Dryopteridaceae)

Debates on speciation processes in pteridophytes have revived. In order to study the evolutionary origin of an apomictic fern species, we investigated the genetic variation in the strictly agamosporous Dryopteris remota. We determined the genotypes of 22 individuals from many different locations within the species' European distribution and of 20 individuals from a Swiss population. A previous study on isozyme variation showed no intraspecific genetic variation in a similar sample set (Schneller and Holderegger, 1994, American Fern Journal 84: 94-98). In contrast to this, four out of 12 random amplified polymorphic DNA (RAPD) primers tested revealed low genetic diversity among individuals of D. remota from different locations. Intrapopulational genetic variation was also very low, but in the single population studied, a unique multiband genotype could be detected. The geographic distribution of genetic variation found in D. remota was best explained by the assumption of a single origin, the accumulation of somatic mutations during spread, and occasional, but effective, events of dispersal over large distances. The present study thus stresses the importance of long-distance dispersal in evolutionary processes and biogeography of ferns.     

Cross-priming of microsatellite loci in subfamily cyprininae (family Cyprinidae): their utility in finding markers for population genetic analysis in three Indian major carps

This study is aimed to identify polymorphic microsatellite markers and establish their potential for population genetics studies in three carp (family cyprinidae; subfamily cyprininae) species, Labeo rohita, Catla catla and Cirrhinus mrigala through use of cyprinid primers. These species have high commercial value and knowledge of genetic variation is important for management of farmed and wild populations. We tested 108 microsatellite primers from 11 species belonging to three different cyprinid subfamilies, Cyprininae, Barbinae and Leuciscinae out of which 63 primers (58.33 %) successfully amplified orthologous loci in three focal species. Forty-two loci generated from 29 primers were polymorphic in these three carp species. Sequencing of amplified product confirmed the presence of SSRs in these 42 loci and orthologous nature of the loci. To validate potential of these 42 polymorphic loci in determining the genetic variation, we analyzed 486 samples of three focal species collected from Indus, Ganges and Brahmaputra river systems. Results indicated significant genetic variation, with mean number of alleles per locus ranging from 6.80 to 14.40 and observed heterozygosity ranging from 0.50 to 0.74 in the three focal species. Highly significant (P < 0.00001) allelic homogeneity values revealed that the identified loci can be efficiently used in population genetics analysis of these carp species. Further, thirty-two loci from 19 primers were useful for genotyping in more than one species. The data from the present study was compiled with cross-species amplification data from previous results on eight species of subfamily cyprininae to compare cross-transferability of microsatellite loci. It was revealed that out of 226 heterologous loci amplified, 152 loci that originated from 77 loci exhibited polymorphism and 45 primers were of multispecies utility, common for 2–7 species.     

Genetic variation within and between Iliamna corei and I. remota (Malvaceae): implications for species delimitation

Iliamna corei and I. remota are classified as endangered species; however, their designation as separate species has been questioned. In order to address this problem, intersimple sequence repeat (ISSR) data were generated to examine patterns of genetic differentiation within and between these two taxa. ISSRs were used to screen individuals for genetic diversity of I. corei from the single known natural population and two garden populations, and individuals of I. remota from the six natural populations and four garden populations. Using ten primers, 140 informative markers were generated. Ninety-four percent of loci detected revealed polymorphisms. Cluster analysis (neighbour-joining, NJ) revealed that the two species are genetically distinct and that the Illinois populations of I. remota were genetically distinct from the Virginia populations of I. remota . Ordination by Principal Coordinate Analysis (PCoA) supported the findings of the NJ, with a separation of I. corei and I. remota . Analysis of Molecular Variance (AMOVA) revealed that the majority of variation detected was within populations, which is consistent with other self-incompatible plants. The results indicate a correlation between the geographical distributions of the species and gene flow.  © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society , 2006, 151 , 345–354.     

Microsatellite primers in the white proteas (Protea section Exsertae, Proteaceae), a rapidly radiating lineage

? Premise of the study: Microsatellite primers were developed in the South African sclerophyllous shrub Protea punctata to investigate the degree of population differentiation within and between P. punctata and closely related species. ? Methods and Results: 10 primer pairs were identified from three individuals of Protea punctata. The primers amplified di- and tri-nucleotide repeats. Across all P. punctata samples, the loci have 8-49 alleles. All primers also amplified in Protea section Exsertae (P. aurea, P. aurea subsp. potbergensis, P. mundii, P. venusta, P. lacticolor, and P. subvestita). The loci had 14-69 alleles across the subgenus. ? Conclusions: These results show the broad utility of microsatellite loci for future studies of population genetics in the white proteas and their potential utility across the entire genus.     

Isolation and characterization of ten microsatellite loci in stingless bee Trigona spinipes (Apidae: Meliponini)

Stingless bees are the most abundant pollinators of Brazilian tropical flora. Trigona spinipes has some of the largest colonies of any stingless bee species found in several types of environment. This work describes the isolation and characterization of microsatellite loci for this species. A microsatellite‐enriched genomic library was constructed and ten primer pairs were designed for T. spinipes. The primers were tested in 20 unrelated individuals. The mean number of alleles was 8.10 and mean observed and expected heterozygosity were 0.655 and 0.680, respectively. Primers were also tested in cross‐species amplification and five loci were successfully amplified in Trigona chanchamayoensis, Trigona hyalinata, Tetragonisca angustula, Partamona mulata and Frieseomelitta varia. The microsatellite primers described herein will be useful for evaluating genetic variability and gaining a better understanding of the population structure of T. spinipes as well as other species of stingless bees.     

Single primer-based DNA amplification as a suitable and low-cost tool for assessing genetic diversity in mangrove crabs

We used single primer-based DNA markers to assess genetic variability of the mangrove crab, Ucides cordatus, collected from four different localities from Pará to Santa Catarina States in Brazil (almost 5000 km distant). Five primers were chosen based on the consistency of the amplified bands and the polymorphism of each locus. A total of 78 loci were amplified in 76 samples; high polymorphism rates were detected in the entire sample (80.8%) and within each locality (73.5-79.5%). Analysis of molecular variance demonstrates significant differences between localities (P < 0.001); however, the Φ(ST) value (0.078) indicates a low level of genetic differentiation, which suggests that U. cordatus larvae can spread over large distances. The variation was distributed among the samples, and most of it was attributed to differences among individuals within localities. Cluster analysis, based on the Jaccard similarity coefficient, and the Mantel test gave similar results to the analysis of molecular variance data. Despite the low level of population structuring, these markers could be used for studying U. cordatus diversity, due to the high level of polymorphism.     

Eight polymorphic microsatellite markers for kelp bass,Paralabax clathratus,amplified in three multiplex polymerase chain reaction sets

Kelp bass, Paralabrax clathratus, is an important sport fishery of California, USA and Baja, Mexico. Here we describe eight microsatellite loci developed for this species. Two loci were derived from known primers for other species in the family, Serranidae and six were developed anew using a clone enrichment protocol. Loci were arranged into three multiplex PCR (polymerase chain reaction) sets for fast throughput; 564 individuals from nine populations across the species’ range were genotyped. Polymorphism ranged from four to 47 alleles and all populations at all loci displayed Hardy–Weinberg and linkage equilibrium.     

Characterization of microsatellite loci in the Jamaican fruit‐eating bat Artibeus jamaicensis and cross‐species amplification

Artibeus jamaicensis is one of the most common bat species in the neotropics, with a well‐defined polygynous social structure in caves. In order to study behaviour and to examine patterns of paternity and relatedness between different harem groups, we developed 14 microsatellite loci from two different enriched genomic libraries. We screened 125 individuals from two different bat colonies and found that polymorphism ranged from five to 13 alleles. Heterozygosity ranged from 63 to 95%. The primers amplified across 14 bat species, indicating their potential utility for population‐level studies in several closely related bat species.     

Very low mitochondrial variability in a stingless bee endemic to cerrado

Partamona mulata is a stingless bee species endemic to cerrado, a severely threatened phytogeographical domain. Clearing for pasture without proper soil treatment in the cerrado facilitates the proliferation of termite ground nests, which are the nesting sites for P. mulata. The genetic consequences of these changes in the cerrado environment for bee populations are still understudied. In this work, we analyzed the genetic diversity of 48 colonies of P. mulata collected throughout the species’ distribution range by sequencing two mitochondrial genes, cytochrome oxidase I and cytochrome B. A very low polymorphism rate was observed when compared to another Partamona species from the Atlantic forest. Exclusive haplotypes were observed in two of the five areas sampled. The sharing of two haplotypes between collection sites separated by a distance greater than the flight range of queens indicates an ancient distribution for these haplotypes. The low haplotype and nucleotide diversity observed here suggests that P. mulata is either a young species or one that has been through population bottlenecks. Locally predominant and exclusive haplotypes (H2 and H4) may have been derived from local remnants through cerrado deforestation and the expansion of a few colonies with abundant nesting sites.     

Trinucleotide Microsatellite Loci and Increased Heterozygosity in Cross-Species Applications in the Social Wasp, Polistes

Though microsatellite loci are usually found to be most polymorphic in the species in which they are first identified, we have found significant increases in polymorphisms in some cross-species applications. We present eight new trinucleotide microsatellite loci derived from two species of social wasps, Polistes annularis and Polistes bellicosus. We assessed the primers designed from these species and the degree of polymorphism in two additional species, P. dorsalis, which is very closely related to P. bellicosus, and P. dominulus, which is an Old World congener, thought to have diverged from New World Polistes over 80 million years ago. Cross-species applications for these microsatellite loci indicate that the priming sites from P. bellicosus loci are conserved in P. dorsalis and amplified similarly sized products with higher heterozygosities than the original species in two of three cases. A locus that was monomorphic in P. annularis had a heterozygosity of 1.0 in the distantly related P. dominulus. Cross-species applications of these loci indicated that alleles were generally of similar lengths in the new and original species when they retained their heterozygosity.     

凉水国家自然保护区天然红松林遗传变异的RAPD分析

采用随机扩增多态DNA(random amplified polymorphic DNA RAPD)技术,研究了凉水国家自然保护区天然红松林的遗传变异水平及其分布规律。16个10bp长度的随机引物在8个实验样地内72个个体中共检测了96个位点,其中55个是多态位点。在种水平上,红松的多态位点比率为P=58.51%;Nei's遗传变异指数为H=0.2868;Shannon's指数为I=0.3654,所有的变异中,有92.47%的变异存在于样地内, 7.53%的变异存在于样地间。根据各遗传多样性在不同样地的分布,初步探讨了红松遗传变异水平与其种群发生及生境的相互关系。     

华东竹黄菌不同居群遗传分化的RAPD分析

为了解竹黄地理居群间的遗传分化,本研究采用随机引物扩增多态性DNA分子标记技术对江苏、安徽和浙江3省的8个居群共32个竹黄样本进行了遗传多样性分析.从50个RAPD引物中筛选得到了5个随机引物,对供试材料的DNA进行扩增,共检测出77个位点,其中多态性位点52个,多态性位点比率为67.53%.UPGMA聚类分析结果表明,这8个居群分为三类:安吉居群、临安居群、宜兴居群、广德居群和泾县居群聚为一类;宁国居群和休宁居群聚为一类;淳安居群单独为一类.遗传多样性分析表明8个竹黄居群中,淳安居群的遗传多样性水平最高,安吉居群的遗传多样性水平最低.Nei's基因多样性指数和Shannon信息指数均表明竹黄物种水平的遗传多样性高于居群水平.     

Isolation of ten tetranucleotide microsatellite loci in the Northern Wheatear (Oenanthe oenanthe)

We isolated 10 polymorphic microsatellite DNA loci from the Northern Wheatear (Oenanthe oenanthe) and optimized these for future studies in population genetics and behavioural ecology. The loci were screened for polymorphism using 107 individuals from one population in Germany. The primers amplified loci with high numbers of alleles ranging from two to 20 alleles per locus.     

Isolation of tetranucleotide microsatellite loci in the burrowing parrot (Cyanoliseus patagonus)

We isolated seven novel polymorphic microsatellite DNA loci from the burrowing parrot (Cyanoliseus patagonus) and optimised them for future studies of population differentiation and genetic variation. The loci were screened for polymorphism using 38 samples from wild individuals from three neighbouring colonies in Argentina. The primers amplified highly variable loci characterised by 3–10 alleles per locus and their observed and expected heterozygosities ranged from 0.15 to 0.78 and 0.15 to 0.81, respectively. When we analysed 52 samples across Argentina and Chile, we found strong genetic differentiation between the Chilean and the Argentinean subspecies as well as significant differentiation between two geographically separated subspecies within Argentina. Our results indicate the suitability of these microsatellites for investigating further questions regarding the population genetics in this species.     

Anonymous nuclear markers for the eastern fence lizard,Sceloporus undulatus

We present results from a screen for de novo variable nuclear loci using a genomic library approach in Sceloporus undulatus, the eastern fence lizard. We tested amplification success for 77 primer pairs in S. undulatus, Sceloporus occidentalis and Sceloporus grammicus. Many loci amplified in all three species suggesting that our primers will be useful for developing sequencing or single nucleotide polymorphism (SNP) genotyping markers in other sceloporine lizards. We also sequenced 19 loci, containing 158 variable sites, for 91 S. undulatus individuals. We report high levels of nucleotide variation in this species with an average of 38 SNPs per kilobase.     

Development of twelve polymorphic microsatellite loci in polyploid endangered Omphalogramma vincaeflora Franch. (Primulaceae)

A total of twelve polymorphic microsatellite loci were developed from polyploid endangered species, Omphalogramma vincaeflora (Primulaceae). These loci were screened for variability among 45 individuals from three populations in China. The primers amplified loci with allele number ranging from 3 to 9, with an average of 4.25 per locus. Polymorphism information content ranged from 0.23 to 0.86. Nei’s genetic diversity ranged from 0.34 to 0.86. These primers provide an opportunity to use polymorphic DNA markers to study the population genetic structure and its breeding system in this species.     

Conservation of (GA)n microsatellite loci between Quercus species

Microsatellite (simple sequence repeat, SSR) amplification was performed in eight different members of the Fagaceae family by using sets of primers developed from sessile oak, Quercus petraea . In total, 136 cases of heterologous amplification were carried out, and 66% resulted in interpretable amplification products. From these, 12 PCR amplification products were sequenced and all 12 contained a sequence homologous to the original locus from Q. petraea . Although SSR primers worked even across different genera, with increasing evolutionary distance there was a clear tendency for decreasing ability to successfully amplify loci and a decreasing proportion of polymorphism amongst those markers which could be amplified. Two of the loci, ssrQpZAG46 and ssrQpZAG110, were polymorphic in all Quercus species tested. Only at one locus, ssrQpZAG58, a specific PCR product could be amplified in all species analysed. For four loci found in two species, we observed significant interspecies differences in the size range of the amplified alleles. Sequence analysis of two alleles showed that the size differences are not only due to variations in the number of (GA) repeats but also to an insertion of approximately 80 nucleotides in the flanking region. Our findings prove the usefulness of SSR markers within and amongst closely related genera of plants.     

Development and characterization of microsatellite markers for the walking goby (Scartelaos viridis; Gobiidae)

Scartelaos viridis (walking goby) is a small edible fish that inhabits warm inshore environments. To provide molecular information of S. viridis, we developed and characterized microsatellite markers for this species. Using (CA)15-enriched genomic libraries of Scartelaos viridis, 44 positive clones were sequenced; 34 sequences contained multiple repeat motifs (di-, tri- and tetra-nucleotide). In all, 23 primer pairs were designed and 15 were successfully amplified. Forty-two S. viridis individuals collected from the East China Sea were used to characterize the polymorphism at each locus. Three loci (13%) were polymorphic, with three to six alleles. The observed and expected heterozygosity ranged from 0.1000 to 0.4500 and from 0.4487 to 0.7580, respectively. The polymorphism information content per locus ranged from 0.4214 to 0.7510. Three loci significantly deviated from the Hardy-Weinberg equilibrium (adjusted P value=0.017); the pairwise tests for linkage disequilibrium between Scvi-1-13 and Scvi-2-11 were significant (P<0.05, adjusted P value=0.017). The low number of polymorphic microsatellite loci may be due to the close genetic relationship of the individuals that we collected and the large size of the motifs.     

Genetic variability and structure of Quercus brantii assessed by ISSR,IRAP and SCoT markers

Persian oak (Quercus brantii Lindl.) is one of the most important woody species of the Zagros forests in Iran. Three molecular marker techniques: start codon targeted (SCoT), inter-simple sequence repeat (ISSR) and inter-retrotransposon amplified polymorphism (IRAP) markers were compared for fingerprinting of 125 individuals of this species collected from different geographical locations of north-west of Iran. A total of 233 bands were amplified by 18 ISSR primers, of which 224 (96.10%) were polymorphic, and 126 polymorphic bands (97.65%) were observed in 129 bands amplified by 10 IRAP primers. Besides, 118 bands were observed for all 10 SCoT primers, of which 113 were polymorphic (95.71%). Average polymorphism information content (PIC) for ISSR, IRAP and SCoT markers was 0.30, 0.32 and 0.38, respectively, and this revealed that SCoT markers were more informative than IRAP and ISSR for the assessment of diversity among individuals. Based on the three different molecular types, cluster analysis revealed that 125 individuals taken for the analysis can be divided into three distinct clusters. The Jaccard's genetic similarity based on the combined data ranged from 0.23 to 0.76. These results suggest that efficiency of SCoT, IRAP and ISSR markers was relatively the same in fingerprinting of individuals. All molecular marker types revealed a low genetic differentiation among populations, indicating the possibility of gene flow between the studied populations. These results have an important implication for Persian oak (Q. brantii) germplasm characterization, improvement, and conservation.