Isolation and characterization of 12 tetranucleotide repeat microsatellite loci from the green‐backed tit (Parus monticolus)

From a partial genomic library enriched for GATA short tandem repeats, we developed 12 polymorphic microsatellite loci from the green‐backed tit (Parus monticolus). We characterized these loci by genotyping 30 adult individuals with unknown relationship. The number of alleles ranged from four to 17 per locus (mean = 9.3 alleles) and the observed heterozygosity for each locus ranged from 0.633 to 0.933 (mean = 0.789). All loci conformed to Hardy–Weinberg expectations. Four of 66 possible pairwise comparisons between loci showed significant gametic disequilibrium.     

Isolation of microsatellites from two species of scleractinian coral

Microsatellites were isolated from two broadcast‐spawning species of scleractinian coral (Platygyra daedalea and Goniastrea favulus) from Australia's Great Barrier Reef. We found 27 microsatellites across both species, although only five loci were polymorphic in each species. Microsatellite loci displayed a wide range of diversity levels with four to 11 alleles per locus (H_O = 0.26–0.91) in P. daedalea and two to seven alleles per locus (H_O = 0.16–0.96) in G. favulus. Most loci showed departures from Hardy–Weinberg equilibrium which may reflect nonrandom mating but may also be related to difficulties associated with coral DNA.     

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.     

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.     

Identification of one X‐linked and five autosomal microsatellite loci in Carabus violaceus (Coleoptera,Carabidae) and their applicability to related taxa

We characterize six microsatellite loci from the ground beetle Carabus violaceus. The polymorphism of the loci was assessed in 46 individuals from one population. The number of alleles ranges between three and eight, and the observed heterozygosities between 0.46 and 0.76. All males are homozygous for locus CVI08106CMPG, whereas most females are heterozygous. This suggests that this locus is situated on the X chromosome. Tests in eight different species of ground beetles showed that four loci could be amplified in C. coriaceus and one each in C. glabratus, C. nemoralis, C. problematicus and Abax parallelepipedus.     

Microsatellite loci in the house fly,Musca domestica L. (Diptera: Muscidae)

Genomic libraries from house flies enriched for (CA)₁₅ and (CAG)₁₀ repeats were constructed by using biotinylated probes. Twenty‐five loci were isolated and evaluated for polymorphisms in wild flies representing two geographically diverse populations. Fourteen of 19 dinucleotide loci, and one of six trinucleotide loci were polymorphic. One hundred and twenty‐seven alleles were detected, 39 of which were private. Average number of alleles per polymorphic locus was 8.4 ± 2.5 and average heterozygosity was 72 ± 4%. F_ST by the private allele method was 0.73. Three of 15 loci showed significant heterozygote deficiencies, attributed to null alleles. Five of 15 loci were amplified in the face fly, Musca autumnalis.     

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).     

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.     

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.     

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.     

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.     

Polymorphic CA/GT and GA/CT microsatellite loci for Ostrinia nubilalis (Lepidoptera: Crambidae)

Ten polymorphic dinucleotide (CA/GT and GA/CT) microsatellite loci suitable for population genetic screening were characterized from enriched partial Ostrinia nubilalis genomic libraries. Sequence from 126 enriched small insert genomic library clones identified 25 CA/GT and 58 GA/CT loci that were unique. Perfect repeats tended to be short (n = 10–12). Ten microsatellites, PCR amplified from a Crawfordsville Iowa population showed a mean of 10 alleles per locus (range six to 20), and six of 10 loci showed heterozygote deficiency. Amplification of eight loci was observed in the sister species O. furnicalis.     

Characterization of polymorphic tetranucleotide microsatellite loci from the Siberian weasel (Mustela sibirica)

Nine tetrarepeat microsatellite loci for Siberian weasel (Mustela sibirica) were characterized. By using 35 individuals of unknown relationship collected from Taiwan, we evaluated the polymorphism of these loci. The number of alleles for each locus ranged from five to 12 (mean = 8.78 alleles) and the observed heterozygosity from 0.429 to 0.962 (mean = 0.688) and all loci conformed to Hardy–Weinberg expectation.     

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.     

Isolation and characterization of eight microsatellite loci from the endangered plant species Hypochaeris salzmanniana (Asteraceae)

We report the isolation and characterization of eight microsatellite loci for Hypochaeris salzmanniana, an endangered species endemic to the southwestern coast of Spain and the Atlantic coast of Morocco. A total of 32 alleles were detected across a sample of 45 individuals, with an average number of 4.0 alleles per locus. The average polymorphic information content (PIC) was 0.533 and the observed (H _O) and expected (H _E) heterozygosity values varied from 0.022 to 0.978 and from 0.434 to 0.759, respectively. Five loci exhibited significant deviation from Hardy–Weinberg equilibrium (P ≤ 0.001) and three pairs of loci showed significant linkage disequilibrium (P ≤ 0.01). The eight loci were tested for transferability in three others species (H. arachnoidea, H. glabra, and H. radicata) belonging to the same section of H. salzmanniana. With the exception of locus Hsalz7, all loci successfully amplified in the three species. These preliminary data confirm the usefulness of microsatellite markers for assessing the ecology and genetic structure of H. salzmanniana and to understand the evolution of species within the section Hypochaeris.     

Genetic diversity and molecular differentiation of Chinese toad based on microsatellite markers

Genetic diversity and population structure of 9 populations of Bufo gargarizans with total 111 samples in China were assessed using seven microsatellite loci. The analysed microsatellite markers produced 161 alleles, varied from 9 to 38 alleles each locus. The number of alleles per population per locus ranged from 4.43 to 10.29. Polymorphic information content showed that all seven loci were highly informative (mean = 0.810 ± 0.071). The average observed heterozygosity was less than the expected (0.353 ± 0.051 and 0.828 ± 0.067, respectively). All tested populations gave significant departures from Hardy–Weinberg equilibrium. Genetic differentiation among the populations was considerably high with the overall and pairwise F _ST values (mean = 0.160 ± 0.039), and showed fairly high level of inbreeding (indicated by a mean F _IS value of 0.504 ± 0.051) and global heterozygote deficit. In comparison to other amphibian studies; however, our results suggested that the level of genetic structuring in B. gargarizans was relatively low in the geographical scale of the study area. Interestingly, the speculated population bottleneck was found to be absent and the analyses provide only weak evidence for a recent contraction in size even though there was severe inbreeding (indicated by the F _IS value) in the Chinese toad populations.     

Ten new highly polymorphic microsatellite loci in the blood clam Scapharca broughtonii

Ten novel microsatellite loci were isolated from blood clam Scapharca broughtonii, and the polymorphisms were examined to estimate genetic variability. The genetic variabilities varied depending on the locus. The number of alleles ranged from 11 to 23, and the observed and expected heterozygosities ranged from 0.63 to 0.93 and 0.66 to 0.95, respectively. Four loci showed significant Hardy–Weinberg disequilibrium at P < 0.05 level. The high variabilities revealed in this study suggest that microsatellites should prove useful for various genetic investigations.     

Isolation and characterization of polymorphic microsatellite loci from a dinucleotide-enriched genomic library of spotted maigre (Nibea albiflora)

In the present study, we reported 13 polymorphic microsatellite loci isolated from a dinucleotide-enriched genomic library of Nibea albiflora. The number of alleles, observed and expected heterozygosity per locus in 44 individuals ranged from 2 to 13, from 0.0909 to 0.9773 and from 0.0886 to 0.9073, respectively. Three loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction. Analysis of linkage disequilibrium showed non-significant among the two pairs of loci. As a result, 13 microsatellite loci probably located on different chromosome pairs and these polymorphic microsatellite loci should provide sufficient level of genetic diversity to investigate the fine-scale population structure and evaluate the breeding strategy in Nibea albiflora. Shichao Xing, Changwei Shao contributed equally to this work.     

Development of microsatellite markers for Polemonium kiushianum (Polemoniaceae), a critically endangered grassland plant species in Japan

Eight microsatellite loci were isolated and characterized for Polemonium kiushianum (Polemoniaceae), an endangered perennial herb endemic to the Kyushu region in Japan. The number of individuals of this species is presumed to be about 400 individuals. The loci showed polymorphisms with two to ten alleles per locus. The expected and observed heterozygositis ranged from 0.039 to 0.744 and 0.039 to 0.784, respectively. Although significant deviations (P < 0.05) from Hardy–Weinberg equilibrium were detected for one locus, no significant deviations were detected for other seven loci. There was no evidence of significant linkage disequilibrium between any two of the loci. The markers described here will be useful to establish conservation programs based on genetic traits of P. kiushianum.     

Microsatellite DNA loci in the rock bream Oplegnathus fasciatus

Nine novel microsatellite loci were isolated from Oplegnathus fasciatus by screening an enriched genomic library using nonradioactive PCR (polymerase chain reaction) techniques. All loci were found to be polymorphic with an average of 8.1 alleles per locus (range 3–15). The mean observed and expected heterozygosities were 0.71 (range 0.40–1.00) and 0.74 (range 0.50–0.90), respectively. Two 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 O. fasciatus.