Variable microsatellite loci isolated from the azure damselfly,Coenagrion puella (L.) (Zygoptera; Coenagrionidae)

We isolated and characterized 10 polymorphic microsatellite loci from the azure damselfly Coenagrion puella (Zygoptera; Coenagrionidae) as part of a study assessing reproductive success and genetic structure in an isolated population of this species. Levels of genetic diversity were assessed in 50 individuals collected from Queen Elizabeth Country Park, Hampshire, UK. The number of alleles per microsatellite loci ranged from three to 22 and the observed and expected heterozygosities varied between 0.26 and 0.84 and between 0.23 and 0.91, respectively. Two loci showed significant (P < 0.05) heterozygote deficits, likely because of null (non‐amplifying) alleles; one pair of loci was in linkage disequilibrium.     

Characterization of eight polymorphic microsatellite markers in the tarnished plant bug,Lygus lineolaris (Palisot de Beauvois)

A partial genomic library of the tarnished plant bug, Lygus lineolaris, enriched for microsatellite sequences was screened to identify marker loci. Eight polymorphic loci suitable for population genetic studies were identified by screening 192 field‐collected insects. The observed number of alleles ranged from four to 21 with an average of 12.25 (SE ± 1.94) while the effective number of alleles ranged from 1.23 to 11.05 with an average of 4.49 (SE ± 1.15). No linkage disequilibria or significant deviations from Hardy–Weinberg expectations were detected at any of the loci. Seven of the eight L. lineolaris microsatellite loci were transferable to Lygus hesperus.     

Development of eighteen polymorphic microsatellite markers in <Emphasis Type="Italic">Scylla paramamosain</Emphasis> by 5′ anchored PCR technique

The mud crab Scylla paramamosain plays a significant role in fishery resources in China. In this study, we developed 18 polymorphic microsatellite markers in this important crab by 5′ anchored PCR technique. A total of 125 alleles were detected in a single population of 32 individuals of S. paramamosain. The number of alleles per locus ranged from five to nine, with the allele size ranging from 166 to 316 bp. The polymorphism information content (PIC), observed and expected heterozygosity ranged from 0.39 to 0.88, from 0.33 to 0.92 and from 0.42 to 0.86, respectively. Three loci (Scypa13, Scypa14 and Scypa15) deviated significantly from Hardy–Weinberg equilibrium (HWE) after Bonferroni correction (P < 0.0028), and no linkage disequilibrium was found between loci pairs. These polymorphic microsatellite markers will be useful for the study of population genetic structure, construction of genetic linkage maps and mapping of economically quantitative trait loci (QTL) in S. paramamosain.     

Microsatellite loci for the weaver ant Oecophylla smaragdina

We developed 13 microsatellite loci in Oecophylla smaragdina from random amplified polymorphic DNA (RAPD) fragments. These loci showed two to 14 alleles in O. smaragdina with expected heterozygosity of each locus from 0.10 to 0.89, and six were also polymorphic in the related species, O. longinoda. The results suggested that the loci will be useful to analyse the genetic structure of Oecophylla species at both the colony and population levels.     

Isolation and characterization of polymorphic microsatellite loci from Yellowcheek (Elopichthys bambusa)

Nine yellowcheek (Elopichthys bambusa) microsatellite loci were isolated using the fast isolation by AFLP of sequences containing repeats (FIASCO) protocol. Three to eight alleles per locus were detected in 29 samples collected from five populations of E. bambusa. The mean number of alleles was 5.6 ± 1.9 and the level of observed heterozygosities ranged from 0.415 to 0.843. These are the first microsatellite loci characterized from E. bambusa that can be used for estimating genetic diversity, population structure and parentage analysis.     

Isolation and characterization of 31 polymorphic microsatellite markers in barfin flounder (Verasper moseri) and the cross-species amplification in spotted halibut (Verasper variegatus)

The construction of a genetic linkage map and evaluation of population genetic diversity both require large numbers of polymorphic molecular markers. In the present study, we report 31 polymorphic microsatellite markers, of which 28 were isolated from two repeat-enriched libraries constructed from genomic DNA, and three were detected in two genes (MCH-R1 and MCH-R2) of barfin flounder (Verasper moseri). A total of 94 alleles were detected with an average of 3.0 alleles per locus. The number of alleles, observed and expected heterozygosity per locus ranged from two to six, from 0.30 to 1.00 and from 0.33 to 0.78, respectively. Three loci significantly deviated from Hardy-Weinberg equilibrium after Bonferroni correction (P < 0.0016) and no significant linkage disequilibrum between pairs of loci was found. Cross-species amplification of these markers was evaluated using the closely related species spotted halibut (Verasper variegatus). This study will potentially be useful for stock management, constructing of a genetic linkage map, mapping economically important quantitative trait loci (QTL), and studying the population genetic diversity of barfin flounder (Verasper moseri).     

Microsatellite loci for paternity assessment in the black swan (Cygnus atratus: Aves)

Seven polymorphic tetranucleotide microsatellite loci were isolated and characterized from the black swan Cygnus atratus Latham, using nonradioactive PCR‐based techniques to screen an enriched genomic library. These loci were highly variable (mean H _E  = 0.75, mean number of alleles = 9.1), and showed no evidence of linkage disequilibrium or null alleles. This locus set is suitable for detecting extra‐pair copulations (combined exclusion probability = 0.999) and for other population genetic applications.     

Isolation and characterisation of di and tri nucleotide microsatellite loci in Rosmarinus officinalis (Lamiaceae), using enriched genomic libraries

An enrichment protocol was used to isolate and characterise microsatellite loci in Rosmarinus officinalis, a Mediterranean chamephyte. Twelve microsatellite loci were characterised and amplified a total of 117 alleles in a sample of 30 individuals from one population, with an average of 9.75 alleles per locus. Observed heterozygosities ranged from 0.333 to 0.900. Cross-species transferability was also assayed in the two other species of the genus. The cumulated probabilities of exclusion for paternity and parentage of the 12 loci were of 0.999971 and 1, respectively, supporting the usefulness of these microsatellite loci for parentage analyses. Nine out of 12 microsatellite loci amplified in the two species and were polymorphic detecting a total of 49 and 45 in R. eriocalyx and R. tomentosus, respectively. Twenty-two alleles were exclusive of R. eriocalyx and 12 of R. tomentosus, additionally, three alleles were shared between these two species but were otherwise absent in the analysed individuals of R. officinalis. In total, this set of markers amplified 154 different microsatellite alleles, supporting their usefulness to conduct population genetic, reproductive biology and hybridisation studies in Rosmarinus.     

Characteristics of 13 polymorphic microsatellite markers in the corn earworm,Helicoverpa zea (Lepidoptera: Noctuidae)

Thirteen polymorphic microsatellite loci suitable for population genetic studies of Helicoverpa zea were discovered by screening partial genomic libraries enriched for microsatellite sequences. Insects collected (N = 96) in Stoneville, Mississippi were used to characterize these markers. The observed and effective number of alleles per locus ranged from two to nine (average 4.46) and from 1.07 to 2.45 (average 1.81), respectively. Fisher exact tests detected significant deviations from Hardy–Weinberg equilibrium at three loci, probably due to inbreeding, null alleles, or Wahlund's effect. Significant genotypic disequilibrium was not detected between any pair of loci.     

Isolation and characterization of 21 microsatellite loci in Lycium chinense and cross-amplification in Lycium barbarum

The present study reports isolation and characterization of 21 polymorphic microsatellite markers developed from Lycium chinense Mill. These markers produced a total of 86 alleles across 30 L. chinense accessions, with an average of 4.1 alleles per locus. Values for observed heterozygosity and polymorphism information content ranged from 0.03 to 0.81 (mean = 0.35) and from 0.03 to 0.78 (mean = 0.31), respectively. At the significance threshold (P < 0.05), 12 loci deviated from Hardy–Weinberg equilibrium, whereas significant linkage disequilibrium values were observed between 52 pairs of loci. All loci were successfully amplified for all L. barbarum accessions. These newly developed polymorphic microsatellite markers will be very useful for programming in the genetic conservation and classification of L. chinense and L. barbarum.     

Isolation and characterisation of polymorphic microsatellite markers for the endangered ferreous limpet <Emphasis Type="Italic">Patella ferruginea</Emphasis> (Gastropoda,Patellidae)

The limpet Patella ferruginea is one of the most endangered marine invertebrates on western Mediterranean rocky shores. We have isolated and characterised 11 polymorphic microsatellites markers to provide new tools to investigate genetic structure and gather information necessary for the proper management of this severely threatened species. The number of alleles per locus ranged from 2 to 16 (mean; Na = 8.37) with an average observed heterozygosity of 0.64 (He = 0.66). The levels of polymorphism uncovered at these loci suggest that they should be useful for population genetic studies, parentage analysis and assessment of connectivity among protected areas. None of the pairwaise comparisons among loci showed significant linkage disequilibrium after sequential Bonferroni correction. All but one locus (Pf-31IF1) conformed to HW equilibrium. Further investigation revealed that departures at Pf-31IF1 were not caused by null alleles. Results from cross-species amplifications suggest that some of these loci may also be useful for Patella tenuis (two loci) P. ulyssiponensis (one locus) and P. piperata (three loci).     

Population genetic structure and phylogeography of cyprinid fish, <Emphasis Type="Italic">Labeo dero</Emphasis> (Hamilton, 1822) inferred from allozyme and microsatellite DNA marker analysis

We examined population structure of Labeo dero (Hamilton, 1822) from different riverine locations in India using 10 polymorphic allozyme and eight microsatellite loci. For analysis, 591 different tissue samples were obtained from commercial catches covering a wide geographic range. Allozyme variability (An = 1.28–1.43, Ho = 0.029–0.071) was much lower than for microsatellites (An = 4.625–6.125, Ho = 0.538–0.633). Existence of rare alleles was found at three allozyme (MDH-2*, GPI* and PGDH*) and at two microsatellite loci (R-3* and MFW-15*). Deviation from Hardy–Weinberg equilibrium (P < 0.05, after the critical probability levels were adjusted for sequential Bonferroni adjustment) could be detected at three loci (EST-1*, -2* and XDH*) whereas, after correction for null alleles, two microsatellite loci (MFW-1*,-15*) deviated from HWE in the river Yamuna. Fst for all the samples combined over all allozyme loci was found to be 0.059 suggesting that 5.9% of the total variation was due to genetic differentiation while microsatellite analysis yielded 0.019 which was concordant to mean Rst (0.02). Hierarchical partition of genetic diversity (AMOVA) showed that greater variability (approx. 95%) was due to within population component than between geographical regions. Based on distribution of genetic differentiation detected by both markers, at least five different genetic stocks of L. dero across its natural distribution could be identified. These results are useful for the evaluation and conservation of L. dero in natural water bodies.     

Characterization of ten polymorphic microsatellite markers for the protected Spanish moon moth <Emphasis Type="Italic">Graellsia isabelae</Emphasis> (Lepidoptera: Saturniidae)

Ten polymorphic microsatellite loci were developed for Graellsia isabelae. Polymorphism was assessed for 20 individuals from a Spanish population (Els-Ports-de-Beseit, Catalonia) and 39 more individuals from one population in the French Alps and six other Spanish localities. Overall, the number of alleles per locus ranged from 5 to 24. Els-Ports-de-Beseit showed an average number of alleles per locus of 9.80 (SD = 4.32), observed heterozygosity was 0.71 (SD = 0.226), and expected heterozygosity was 0.788 (SD = 0.146). Genotypic frequencies conformed to Hardy–Weinberg equilibrium at the Catalonian population, and no evidence for linkage disequilibrium was observed. Multilocus genotypes resulting from this set of markers will be useful to determine genetic diversity and differentiation within and among populations of this highly protected moth. Several loci amplified and resulted polymorphic in two related species: two loci in Actias neidhoeferi, and three loci in A. luna.     

Development and characterization of microsatellite markers in the fungus Antrodia cinnamomea from dbEST database

We developed and characterized 8 polymorphic microsatellite loci or simple sequence repeats from the expressed sequence tags database of the medicinal fungus, Antrodia cinnamomea. Variability was assessed in a panel of 23 strains collected from various regions of Taiwan. The number of alleles per polymorphic locus ranged from two to seven loci with an average of 4.375 alleles per locus. The observed and expected heterozygosity values ranged from 0.130 to 0.783 and from 0.122 to 0.809, respectively. No significant Hardy–Weinberg equilibrium was detected (P < 0.01) at all loci. The EST-SSR markers developed in the present study can be used for strain identification and population genetic studies in A. cinnamomea.     

Differential introgression from a sister species explains high FST outlier loci within a mussel species

Scanning genomes for loci with high levels of population differentiation has become a standard of population genetics. F_ST outlier loci are most often interpreted as signatures of local selection, but outliers might arise for many other reasons too often left unexplored. Here, we tried to identify further the history and genetic basis underlying strong differentiation at F_ST outlier loci in a marine mussel. A genome scan of genetic differentiation has been conducted between Atlantic and Mediterranean populations of Mytilus galloprovincialis. The differentiation was low overall (F_ST = 0.03), but seven loci (2%) were strong F_ST outliers. We then analysed DNA sequence polymorphism at two outlier loci. The genetic structure proved to be the consequence of differential introgression of alleles from the sister‐hybridizing species Mytilus edulis. Surprisingly, the Mediterranean population was the most introgressed at these two loci, although the contact zone between the two species is nowadays localized along the Atlantic coasts of France and the British Isles. A historical contact between M. edulis and Mediterranean M. galloprovincialis should have happened during glacial periods. It proved difficult to disentangle two hypotheses: (i) introgression was adaptive, implying edulis alleles have been favoured in Mediterranean populations, or (ii) the genetic architecture of the barrier to edulis gene flow is different between the two M. galloprovincialis backgrounds. Five of the seven outliers between M. galloprovincialis populations were also outliers between M. edulis and Atlantic M. galloprovincialis, which would support the latter hypothesis. Differential introgression across semi‐permeable barriers to gene flow is a neglected scenario to interpret outlying loci that may prove more widespread than anticipated.     

Isolation and characterization of microsatellite loci in Tripterygion delaisi

We isolated 49 microsatellite loci from a genomic library of Tripterygion delaisi×anthosoma enriched for CA and GA repeats. Ten loci were screened in 30 individuals with high numbers of alleles per locus (averaging 15.5 ± 2.86) and observed heterozygosity (averaging 0.765 ± 0.052). No deviations from Hardy–Weinberg expectations were detected. These highly polymorphic markers will be useful in determining the spatial patterns of genetic diversity between and within subspecies of Tripterygion delaisi.     

Genetic structure and differentiation in cultivated fig (<Emphasis Type="Italic">Ficus carica</Emphasis> L.)

One hundred ninety-four germplasm accessions of fig representing the four fig types, Common, Smyrna, San Pedro, and Caprifig were analyzed for genetic diversity, structure, and differentiation using genetic polymorphism at 15 microsatellite loci. The collection showed considerable polymorphism with observed number of alleles per locus ranging from four for five different loci, MFC4, LMFC14, LMFC22, LMFC31 and LMFC35 to nine for LMFC30 with an average of 4.9 alleles per locus. Seven of the 15 loci included in the genetic structure analyses exhibited significant deviation from panmixia, of which two showed excess and five showed deficiency of heterozygote. The cluster analysis (CA) revealed ten groups with 32 instances of synonymy among cultivars and groups differed significantly for frequency and composition of alleles for different loci. The principal components analysis (PCA) confirmed the results of CA with some groups more differentiated than the others. Further, the model based Bayesian approach clustering suggested a subtle population structure with mixed ancestry for most figs. The gene diversity analysis indicated that much of the total variation is found within groups (H _G /H _T = 0.853; 85.3%) and the among groups within total component (G _GT = 0.147) accounted for the remaining 14.7%, of which ~64% accounted for among groups within clusters (G _GC = 0.094) and ~36% among clusters (G _CT = 0.053). The analysis of molecular variance (AMOVA) showed approximately similar results with nearly 87% of variation within groups and ~10% among groups within clusters, and ~3% among clusters. Overall, the gene pool of cultivated fig analyzed possesses substantial genetic polymorphism but exhibits narrow differentiation. It is evident that fig accessions from Turkmenistan are somewhat genetically different from the rest of the Mediterranean and the Caucasus figs. The long history of domestication and cultivation with widespread dispersal of cultivars with many synonyms has resulted in a great deal of confusion in the identification and classification of cultivars in fig.     

Genetic variation within and between populations of hermaphroditic <Emphasis Type="Italic">Bulinus truncatus</Emphasis> tetraploid freshwater snails of the Albertine Rift,East Africa

Genetic variability within and among Bulinus truncatus of the Albertine Rift freshwater bodies were assessed to investigate the degree of inbreeding and gene flow in the snail populations. The effect of ploidy on the genetic structuring of B. truncatus is also described. We characterized the genetic structure of seven B. truncatus populations from Lake Albert, Lake Kivu, and Katosho swamp in Tanzania using five polymorphic microsatellite loci. Genetic differentiation was quantified using pairwise FST values and Nei’s standard genetic distances. Different alleles were observed across all loci and genetic diversity was low although it varied greatly across populations; observed heterozygosity was, however, higher than the expected heterozygosity in three of the populations studied. Significant heterozygote deficiencies were observed coupled with significant linkage disequilibria in five populations for all the five loci examined in this study. We found significant genetic differentiation among the seven freshwater bodies; private alleles were observed across all loci indicating restricted or absence of gene flow between populations. Limited snail dispersal and the reproductive biology of B. truncatus are the major forces shaping the genetic variation observed. Low genetic variation within B. truncatus populations exposes them to a high parasite infection risk as predicted in the Red Queen hypothesis.     

New microsatellite markers for the snow crab Chionoecetes opilio (Brachyura: Majidae)

Nine novel microsatellite loci were isolated from Chionoecetes opilio by screening an enriched genomic library using nonradioactive polymerase chain reaction techniques, and the polymorphisms were examined to estimate genetic variability. The genetic variabilities varied depending on the locus. All loci were found to be polymorphic with an average of 9.7 alleles per locus (range 3–25). The observed and expected heterozygosities ranged from 0.80 to 0.98 and from 0.56 to 0.95, respectively. Five loci showed significant Hardy–Weinberg disequilibrium at the P < 0.05 level. The high variabilities revealed in this study suggest that these microsatellite loci should provide useful markers for genetic variation monitoring of C. opilio.     

Isolation and characterization of ten novel microsatellite loci in the red-winged tinamou (<Emphasis Type="Italic">Rhynchotus rufescens</Emphasis>, Tinamiformes,Aves) and cross-amplification in other tinamous

We describe the isolation and characterization of ten microsatellite loci from the red-winged tinamou (Rhynchotus rufescens) and also evaluated the cross-amplification of these loci and other ten loci previously developed for the great tinamou (Tinamus major) in other tinamous. Genetic variability was assessed using 24 individuals. Six loci were polymorphic with moderate to high number of alleles per locus (2–12 alleles) and showed expected heterozygosity (HE) ranging from 0.267 to 0.860. All loci conformed to the Hardy–Weinberg expectation and linkage disequilibrium was not significant for any pair of loci. This battery of polymorphic loci showed high paternity exclusion probability (0.986) and low genetic identity probability (4.95 × 10⁻⁵), proving to be helpful for parentage tests and population analyses in the red-winged tinamou. The cross-amplification was moderate where of the 160 locus/taxon combinations, 46 (28.75%) successfully amplified.