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.     

Polymorphic microsatellite DNA markers for Banksia attenuata (Proteaceae)

We developed 11 polymorphic microsatellite DNA markers for an Australian native shrub Banksia attenuata. The number of alleles per locus in 50 individuals varied from five to 18, observed and expected heterozygosities ranged from 0.300 to 0.740 and from 0.537 to 0.918, respectively. Six loci showed no significant deviation from Hardy–Weinberg equilibrium (P > 0.05), and null alleles appear to exist at locus BA‐B1. All loci showed independent inheritance.     

Development of microsatellite marker loci for European hazelnut (<Emphasis Type="Italic">Corylus avellana</Emphasis> L.) from ISSR fragments

Polymorphic microsatellite markers were developed for European hazelnut (Corylus avellana L.) from the sequences of inter-simple sequence repeat (ISSR) fragments and flanking regions. Twenty-five ISSR primers were used to generate fragments for cloning. Of the 520 unique sequences obtained, 41 contained long internal repeats (≥20 bp) with flanking sequences sufficient for primer design. From these, we developed 23 new polymorphic microsatellite loci. The flanking sequences were obtained for fragment ends by chromosome walking, and an additional 47 polymorphic markers were developed. Two additional polymorphic markers were developed from a GA-enriched library. The 72 new marker loci were characterized using 50 diverse hazelnut accessions. For the internal repeat loci, the number of alleles per locus ranged from 2 to 16, with a mean of 7.52. Mean values for expected heterozygosity (H_e), observed heterozygosity (H_o), and polymorphism information content (PIC) were 0.62, 0.59, and 0.58, respectively. The estimated frequency of null alleles (r) was ≥0.05 at six of the 23 loci. For the 47 marker loci developed from fragment ends, the number of alleles per locus ranged from 2 to 16, with a mean of 7.30. Mean values for H_e, H_o, and PIC were 0.62, 0.47, and 0.58, respectively. The estimated frequency of null alleles (r) was ≥0.10 at 18 of the 47 loci. Of the 70 loci developed from ISSR and flanking sequences, 50 segregated in our mapping population and were assigned to linkage groups.     

Characterization of expressed sequence tag (EST)-derived microsatellite loci in the fire ant Solenopsis invicta (Hymenoptera: Formicidae)

The fire ant Solenopsis invicta has received considerable attention as a damaging invasive pest. In the present study, 12 polymorphic microsatellite loci were developed from publicly available expressed sequence tags (ESTs) of S. invicta. The number of alleles varied between 3 and 12, with an average of 6.42 per locus. The observed and expected heterozygosities ranged from 0.4167 to 1.0000 and from 0.3446 to 0.8543, respectively. These informative EST-derived microsatellite loci provide a powerful complementary tool for genetic studies of S. invicta.     

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.     

Identification and characterization of nine polymorphic microsatellite loci in the North American pika,Ochotona princeps

Nine polymorphic microsatellite loci were developed for the North American pika (Ochotona princeps) from di‐ and tetranucleotide repeat‐enriched genomic libraries. Polymorphism was assessed for 165 individuals from eight geographical locations in the western United States. All loci were polymorphic. The number of alleles per locus ranged from three to 14, with observed heterozygosity between 0.189 and 0.822. All loci were in Hardy–Weinberg equilibrium (< 0.05). Regional differences were evident with unique alleles at multiple loci in six of eight populations.     

Microsatellite markers for <Emphasis Type="Italic">Pseudopanax</Emphasis> trees and their cross-species amplification in the Araliaceae

In order to study hybridisation, taxonomic boundaries and phylogeography in the genus Pseudopanax (Araliaceae), we developed seven novel microsatellite loci from enriched genomic libraries of P. lessonii and P. crassifolius. These loci were characterised in 16 individuals from a single population of P. crassifolius, and displayed 2–11 alleles per locus. Observed heterozygosities ranged from 0.063 to 0.688. Most loci were polymorphic in the closely related Pseudopanax species, and several loci amplified widely across the Araliaceae.     

Development of simple sequence repeat markers for the plant pathogenic rust fungus Puccinia triticina

Eighteen polymorphic di‐ and trinucleotide simple sequence repeat markers were developed for the phytopathogenic rust fungus Puccinia triticina. The allelic diversity varied from two to nine alleles per locus. Levels of observed heterozygosity ranged from 0.095 to 0.952. Seven of the loci deviated significantly from Hardy–Weinberg equilibrium (P < 0.002) with 70% having levels of observed heterozygosity higher than expected heterozygosity. Null allele(s) were observed for locus PtSSR76 with a frequency of 9%. A preliminary screen of other cereal rust fungi (P. coronata, P. graminis, P. recondita and P. striiformis) indicated that these primer pairs are specific to P. triticina.     

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.     

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 characterization of polymorphic microsatellite loci in <Emphasis Type="Italic">Achnatherum sibiricum</Emphasis> (Poaceae)

Achnatherum sibiricum is a threaten and toxic perennial bunchgrass mainly in the north of China. We developed 11 polymorphic microsatellite loci from A. sibiricum by combining biotin capture method. After validating and scoring, these loci were polymorphic in a test population with the range of alleles from four to 13 per locus. The observed and expected heterozygosities ranged from 0.1649 to 0.5624 and from 0.3071 to 0.8826, respectively. All 11 microsatellite markers were in Hardy–Weinberg equilibrium. These polymorphic microsatellites will be useful for genetic diversity analysis and population structure construction for A. sibiricum.     

Isolation and characterization of polymorphic microsatellite loci from a dinucleotide-enriched genomic library of starry flounder (Platichthys stellatus) and cross-species amplification

Starry flounder (Platichthys stellatus) is a rare fish species in China. Here, we reported 12 polymorphic microsatellite loci isolated from a dinucleotide-enriched genomic library of starry flounder (P. stellatus). The number of alleles, observed and expected heterozygosity per locus in 30 individuals ranged from two to six, from 0.2500 to 1.0000 and from 0.4512 to 0.7667, respectively. One locus significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction and no significant linkage disequilibrium between pairs of loci was found. Cross-species amplification of these microsatellite loci in additional three fish species was performed. These polymorphic microsatellite loci would be useful for investigating genetic population structure and construction of genetic linkage map in P. stellatus. Guidong Miao and Changwei Shao have contributed equally.     

Development of 15 polymorphic genic microsatellite DNA markers of Pacific abalone Haliotis discus hannai

Fifteen polymorphic genic microsatellite DNA markers were developed based on the expressed sequence tags of Pacific abalone Haliotis discus hannai we ourselves generated, which were used to type 32 individuals representing a cultured population. The number of alleles each locus ranged from two to 12. The observed and expected heterozygosities ranged from 0.094 to 0.969 and from 0.091 to 0.878, respectively. Among 15 loci, four were found to deviate significantly from Hardy–Weinberg equilibrium (P_HW < 0.001). No linkage disequilibrium was found between these loci. It is certain that these markers will facilitate the management and exploitation of the genetic resource of Pacific abalone.     

Microsatellite primers for three Western Atlantic Farfantepenaeus prawn species

Microsatellite loci were identified for three closely related penaeid species, Farfantepenaeus subtilis, F. paulensis and F. sp., from genomic libraries enriched for CA repeats. Seven out of nine highly polymorphic loci detected were amplified across all three species. Between four and 64 alleles were recorded per locus (average = 36). The average observed heterozygosity ranged from 0.094 to 0.897 (mean = 0.613), while the expected heterozygosity ranged from 0.091 to 0.985 (mean = 0.822).     

Microsatellite development for the endangered Yangtze sturgeon (Acipenser dabryanus Duméril, 1869) using 454 sequencing

The Yangtze sturgeon (Acipenser dabryanus) is an endemic species in China. Using 454 sequencing, eight polymorphic tri‐, tetra‐, penta‐, and hexanucleotide microsatellite loci were isolated in this study. The raw sequence data from a one‐eighth run of 454 sequencings were 38.0 Mbp containing 94 222 reads/sequences. Of 80 microsatellite loci, only eight loci were polymorphic in a population of 30 individuals. The number of alleles per locus ranged from 4 to 14 (mean 7.62), and the observed heterozygosities varied between 0.46 and 0.88 (mean 0.74). Cross amplification was tested in congeneric species Acipenser sturio and Acipenser sinensis. These new microsatellite markers will be useful for further studies on genetic variation, parentage analysis, and conservation management for this critically endangered species.     

Microsatellite loci transferability from Theobroma cacao to Theobroma grandiflorum

Theobroma grandiflorum (cupuassu) is an important fruit tree native to the Brazilian Amazon. Forty‐eight microsatellite loci developed for the congener Theobroma cacao were tested in cupuassu, and 29 (60.4%) produced robust alleles. The analyses of 216 cupuassu accessions using the 21 polymorphic microsatellite loci revealed a total of 113 alleles. The number of alleles per polymorphic locus ranged from two to 11, with an average of 5.38 alleles per locus. The average observed heterozygosity was 0.343, while the mean expected heterozygosity was 0.614. The successful transferability of T. cacao microsatellite primers to cupuassu was consistent with currently accepted phylogeny.     

Development and characterization of 10 novel polymorphic microsatellite loci in Homatula potanini (Günther, 1896)

Homatula potanini is a genus Paracobitis which belongs to the family Cobitidae, subfamily Nemacheilinae. We first isolated ten novel microsatellite DNA loci using the FIASCO protocol. The number of observed alleles per locus ranged from 5 to 14. The observed heterozygosity and expected heterozygosity ranged from 0.3750 to 0.9375 and 0.4028 to 0.8819 respectively. The polymorphic information content ranged from 0.3515 to 0.8556 (M = 0.6170). No significant linkage disequilibrium was detected among the loci. In the present study, we have identified 10 new markers for H.  potanini. These loci should provide sufficient levels in the evaluation of genetic diversity and designing effective conservation programs of H.  potanini.     

Isolation and characterization of polymorphic microsatellite loci in the Hawaiian flycatcher,the elepaio (Chasiempis sandwichensis)

Thirteen polymorphic microsatellite loci were isolated and characterized from two clades of an endemic Hawaiian flycatcher, the elepaio (Chasiempis sandwichensis). Seven dinucleotide repeats and one trinucleotide repeat were cloned from Kauai elepaio; five dinucleotide repeats were cloned from Oahu elepaio. Polymorphism was assessed in a sample of Oahu elepaio (n = 22) revealing two to 16 alleles per locus. Observed heterozygosity ranged from 0.14 to 0.91. However, linkage analysis exposed highly significant linkage disequilibrium between two of the most polymorphic loci. Twelve loci are therefore expected to be useful for investigations of population structure.     

Isolation and characterization of 14 polymorphic microsatellite loci in the ark shell Scapharca subcrenata (Bivalvia: Arcidae)

The ark shell Scapharca subcrenata is an important fishery resource, but has been suffering from severe population decline due to the deterioration of the coastal environment and over-exploitation. To investigate the genetic variation and structure among populations of S. subcrenata, 14 polymorphic microsatellite loci were developed. The number of alleles per polymorphic locus ranged from 11 to 29 and the observed and expected heterozygosity varied from 0.400 to 0.923 and from 0.705 to 0.965, respectively. These microsatellites will help advance the investigation of the genetic population structure and genetic diversity in this species.