Isolation and characterization of 19 novel microsatellite loci in Rhododendron aureum and Rhododendron brachycarpum (Ericaceae)

Nineteen polymorphic microsatellite loci are described for Rhododendron aureum Georgi (Ericaceae), an endangered species in Korea, and the closely related Rhododendron brachycarpum. The sequences containing repeat motifs were identified using low-depth next generation sequencing and 148 microsatellite loci were examined for amplification success and the detection of polymorphisms. All 19 loci were polymorphic and the number of alleles for these markers varied from 4 to 25, with an average of 15 alleles per locus. Three R. aureum loci showed departure from Hardy–Weinberg equilibrium (HWE) and one R. brachycarpum locus deviated significantly from HWE. These microsatellite loci will be useful for investigating the genetic diversity and population structure of both species.     

Polymorphic microsatellite loci for species of Australian freshwater cod, <Emphasis Type="Italic">Maccullochella</Emphasis>

The Australian freshwater cod genus, Maccullochella is represented by three species: Murray cod, M. peelii peelii, eastern freshwater cod, M. ikei, and trout cod, M.macquariensis. Seven novel microsatellite loci from M. ikei and six previously published loci from M. peelii peelii were tested on wild populations of Murray, eastern and trout cod. Levels of polymorphism varied between species with 13 loci polymorphic in Murray cod, 9 in trout cod and 7 in eastern cod. Observed heterozygosities ranged from 0.053 to 0.842. This suite of microsatellite loci will facilitate future studies of the genetic status of wild and hatchery bred populations of Maccullochella.     

Isolation and characterization of nine polymorphic microsatellite markers of the marbled rockfish Sebastiscus marmoratus (Scorpaeniformes, Scorpaenidae)

Sebastiscus marmoratus is a typical sedentary fish and usually lives at the bottom of the coastal zone from Japan to the East China Sea. Next-generation sequencing platform was used to obtain microsatellite loci sequences of S. marmoratus, among which forty-eight perfect microsatellite repeats were suitable for primer design. Nine polymorphic microsatellite markers were exploited and characterized for it. The allele number of all polymorphic microsatellite markers ranged from 4 to 12. Expected and observed heterozygosity varied from 0.302 to 0.909 and from 0.190 to 0.625, respectively. In further studies, the genetic variation and population structure of this species would be investigated using these microsatellite loci.     

Characterization of microsatellite loci in village indigobirds Vidua chalybeata and cross‐species amplification in estrildid and ploceid finches

We characterized 11 microsatellite primer pairs for the village indigobird Vidua chalybeata. The loci were highly polymorphic, with 7–13 alleles per locus. Gene diversity, estimated as expected heterozygosity, ranged from 0.52 to 0.86, and was generally matched by levels of observed heterozygosity (0.49–0.91). Many of these primer pairs amplified polymorphic loci in cross‐species amplification trials with a variety of estrildid and ploceid finches and a sparrow, Passer griseus. These primers will be valuable for genetic analyses of the brood parasitic indigobirds and whydahs (genus Vidua) as well as other Old World finches.     

Microsatellite markers for the palaeotropic pioneer tree genus Macaranga (Euphorbiaceae) and their cross‐species transferability

We developed primer sequences for five polymorphic microsatellite loci in the tropical ant‐plant genus Macaranga (Euphorbiaceae). Population genetic parameters were determined on the basis of 30 individuals from each of two Macaranga species in Borneo. Allele numbers per locus ranged from three to 13. Expected and observed heterozygosities ranged from 0.160 to 0.850 and from 0.130 to 0.700, respectively. Four of the five primer pairs cross‐amplify polymorphic PCR products in a wide range of Macaranga species.     

Characterization of 11 novel polymorphic microsatellite loci in the threatened Korean loach,Iksookimia koreensis,isolated using a next-generation sequencing method

To investigate the genetic diversity and population structure of Iksookimia koreensis, we characterized 11 novel polymorphic microsatellite markers developed using next-generation sequencing. The number of alleles per locus ranged from 4 to 10 (average = 6.26). Observed and expected heterozygosities ranged from 0.333 to 0.866 and from 0.375 to 0.866, respectively. No loci showed significant deviation from Hardy–Weinberg equilibrium (HWE). These loci were also used successfully to study the genetic diversity of a closely related species, Iksookimia longicorpa. Four of the 11 loci amplified in the two species showed different allele frequency and distribution, indicating deep genetic divergence between I. koreensis and I. longicorpa. The newly developed microsatellite markers reported here will provide a useful tool for examining gene flow, population genetic structure, and genetic diversity of these species.     

Application of Microsatellite Primers Developed for Polistes in the Independent-Founding Wasp Polists satan Bequaert (Hymenoptera: Vespidae)

Microsatellite primers developed for a given species are sometimes useful for another in the same genus and in other genera within the same family, making possible to search for pre-existing suitable primers in the databanks such as GenBank. We examined whether existing primers developed for Polistes could be used for Polistes satan Bequaert. We tested 50 microsatellite primers from three Polistes species and found that six microsatellite loci show polymorphism in size in P. satan. These six loci were highly polymorphic, having four to 15 alleles in P. satan with an expected heterozygosity of 0.525?C0.832. These loci can be used to study parameters concerning genetic relatedness such as social interactions in colonies and genetic conflicts of interest among nestmate individuals.     

Isolation and characterization of eighteen polymorphic microsatellite loci in Schizopygopsis younghusbandi Regan and cross-amplification in three other schizothoracinae species

Eighteen polymorphic microsatellite loci were isolated from Schizopygopsis younghusbandi Regan and the characterization of these loci was assessed in 46 individuals collected from the Yarlung Tsangpo River in Tibet, China. The number of alleles per locus ranged from 2 to 14. The expected heterozygosity and Shannon-Wiener diversity index ranged from 0.022 to 0.879 and from 0.059 to 2.313, respectively. The cross-species amplification and applicability of these loci were tested in three other Schizothoracinae species belonging to Schizothorax and Oxygymnocypris. These loci will be useful for the evaluation of genetic diversity and population genetic structure in S. younghusbandi and other related species.     

Isolation and characterization of polymorphic microsatellite loci in the field vole,Microtus agrestis,and their cross‐utility in the common vole,Microtus arvalis

We developed nine polymorphic microsatellite loci for the field vole, Microtus agrestis. The number of alleles ranged from five to 15 and observed heterozygosities ranged from 0.40 to 1.00. We also tested the microsatellite loci for amplification and polymorphism in the congeneric species Microtus arvalis. Five of the nine loci were successfully analysed in this species. The microsatellite markers will be employed in studies of reproductive success and fine‐scale spatial genetic structure.     

Isolation and characterization of polymorphic microsatellite loci from a dinucleotide-enriched genomic library of obscure puffer (Takifugu obscurus) and cross-species amplification

Obscure puffer (Takifugu obscurus) is an anadromous fish species in China. Here, we reported 10 polymorphic microsatellite loci isolated from a dinucleotide-enriched genomic library of T. obscurus. The number of alleles, observed and expected heterozygosity per locus in 30 individuals ranged from four to 10, from 0.57 to 0.86 and from 0.68 to 0.90, respectively. Three loci 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 T. obscurus.     

Isolation and characterization of 19 polymorphic microsatellite loci from Neosalanx taihuensis,a rapidly invasive and adaptative species

Neosalanx taihuensis can very rapidly adapt to new environmental conditions and quickly becomes the dominant species when introduced to lakes and reservoirs. To better understand the genetic mechanism responsible for this remarkable ability to invade and adapt to diverse ecosystems, 19 polymorphic microsatellite loci were isolated from the genomic DNA of N. taihuensis. These microsatellite loci were identified from 24 individuals from an invasive population sampled in Fuxian Lake, Yunnan Plateau, Southwest China. The number of alleles per locus ranged from 2 to 14. Observed and expected heterozygosities ranged from 0.0417 to 0.8636 and from 0.3112 to 0.9202, respectively. The polymorphism information content (PIC) varied from 0.2583 to 0.8928. These microsatellite loci will be essential in performing molecular analyses of genetic diversity and genetic structure in the future. This will ultimately facilitate better understanding of the genetic mechanisms that allow N. taihuensis to successfully invade and adapt to new ecosystems.     

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.     

Novel polymorphic microsatellites for the red‐legged partridge (Alectoris rufa) and cross‐species amplification in Alectoris graeca

The red‐legged partridge, Alectoris rufa, is an endemic species of the southwestern Mediterranean, and the most popular game bird in the Iberian Peninsula. A total of 27 microsatellite loci was isolated from an enriched genomic library of A. rufa. Six perfect GT microsatellites were characterized and optimized in 45 individuals of A. rufa. All loci revealed high levels of polymorphism with a number of alleles that ranged from three to 13. Observed heterozygosity ranged from 0.2 to 0.6. Cross‐species amplification showed that all loci were also polymorphic in rock partridge, Alectoris graeca. The new markers will be useful in determining hybridization between both species of Alectoris.     

A Natural Homoploid Hybrid between Centaurea horrida and Centaurea filiformis (Asteraceae) as Revealed by Morphological and Genetic Traits

Studies over the last two decades demonstrate that hybridization has played an integral role in the evolution of several sections of the genus Centaurea. Nevertheless, natural hybridization between narrow Mediterranean endemic Centaurea species has not been documented as yet. A population of fertile Centaurea individuals exhibiting intermediate morphological traits between two Sardinian narrow endemics, C. horrida and C. filiformis, was identified at the Tavolara Islet (Sardinia, Italy). Intermediate leaf length and head width characterized this population, suggesting its hybrid origin. The putative hybrid population was structured (i.e., composed of seedlings, saplings and adult individuals) and had a relatively high levels of seed production. The number of chromosomes was identical to that of the proposed progenitors (2n?=?18). Genotyping at five microsatellite loci showed that the putative hybrid possessed several alleles in common with the proposed parental species and intermediate values of genetic differentiation, as indicated by both F _ST and R _ST, between C. horrida and C. filiformis. We therefore conclude that the studied intermediate population is of hybrid origin, and discuss possible mechanisms of its reproductive isolation from the parental species, potential re-introgression, and evolutionary implications of this hybridization.     

Isolation and characterization of microsatellite loci in the Japanese pipistrelle (Pipistrellus abramus)

We describe the first set of ten microsatellite markers isolated in Pipistrellus abramus. The number of alleles per locus ranged from 7 to 13. The observed and expected heterozygosities values ranged from 0.486 to 0.971 and from 0.752 to 0.876, respectively. Three loci revealed significant departure from Hardy–Weinberg equilibrium and no linkage disequilibrium was found between loci pairs. These informative microsatellite markers will be a powerful molecular tool for studying the population genetic structure of P. abramus, as well as other species of this genus.     

Isolation and characterization of microsatellite markers for the heavily exploited rockfish Sebastes schlegeli, and cross-species amplification in four related Sebastes spp.

Here we report development and characterization of 14 polymorphic microsatellite loci from Sebastes schlegel. Polymorphism at these loci revealed from 3 to 23 alleles. The observed heterozygosity ranged from 0.34 to 1.00, and the expected heterozygosity ranged from 0.31 to 0.95. No linkage disequibrium was found. Two loci were significantly deviated from HWE (P < 0.01). The 14 loci were also surveyed in four other Sebastes species and 12 loci successfully amplified, where allelic diversity ranged from highly polymorphic to monomorphic. These results demonstrate these microsatellite markers can be used for the study of intra- and inter-specific genetic diversity.     

Isolation and characterization of fifteen polymorphic microsatellite loci in the threatened freshwater mussel Solenaia oleivora (Bivalvia: Unionidae)

Freshwater mussels are among the most endangered fauna in the world, while little is known about their genetic diversity. We isolated 15 polymorphic microsatellite loci in a threatened freshwater mussel, Solenaia oleivora. The characteristics of these loci were tested with 46 individuals. The number of alleles per locus ranged from 2 to 15, and the PIC value ranged from 0.250 to 0.880. The observed and expected heterozygosity ranged from 0.167 to 0.929 and from 0.297 to 0.900, with an average of 0.562 and 0.647, respectively. Eight loci deviated significantly from Hardy–Weinberg equilibrium. These polymorphic microsatellites will be useful for the assessment of population genetic diversity and resource conservation of S. oleivora. Cross-species amplification demonstrated that six of 15 microsatellite loci amplified successfully in two other unionid species.     

Isolation and characterization of microsatellite loci in Lesquerella fendleri (Brassicaceae) and cross‐species amplification

Fifteen novel microsatellite primer pairs are presented for Lesquerella fendleri, which were developed from seven dinucleotide, five trinucleotide and three tetranucleotide microsatellite DNA loci. These loci were characterized for 40 individuals from 24 localities throughout the species range. The number of alleles observed per locus ranged from three to 16, the observed heterozygosity ranged from 0.175 to 0.750, and the polymorphic information content ranged from 0.218 to 0.889. Cross‐species transferability tested on nine species of Lesquerella and one species of the related genus Physaria indicates that these primer pairs may be useful for population genetic studies of other species in Lesquerella and possibly other closely related genera.     

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.     

Isolation and Characterization of Twelve Microsatellite Loci for Rockfish (Sebastes)

We describe the first microsatellites for rockfishes in the diverse genus Sebastes. Clones containing microsatellites were isolated from the genomic library of a quillback rockfish, Sebastes maliger. Twelve microsatellites are characterized; six of these are polymorphic in quillback rockfish, and eight are polymorphic in at least one rockfish species on which they were tested. The number of alleles per variable locus ranged from 4 to 15 and averaged 6.8. The expected heterozygosities ranged from 0.38 to 0.79 and averaged 0.60 in these loci. These loci should prove valuable in studies examining species identification, population genetics, hybridization, paternity, kinship, and microsatellite evolution. Received September 8, 1998; accepted November 23, 1998.