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.     

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 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.     

Isolation of eight microsatellites loci from the saddled bream, <Emphasis Type="Italic">Oblada melanura</Emphasis> and cross-species amplification in two sea bream species of the genus <Emphasis Type="Italic">Diplodus</Emphasis>

We have developed eight new microsatellite markers for the saddled bream (Oblada melanura) from an enriched genome library protocol. All these loci are polymorphic, with mean allelic diversity of 14.75 (range 3–22), and expected and observed heterozygosities from 0.233 to 0.918 and 0.212 to 0.913, respectively. Cross-species tests in two close relatives of the genus Diplodus (D. sargus and D. vulgaris) revealed successful amplifications at 6 out of 8 loci, with means allele number of 6.67 (range 4–10) and 6.50 (range 4–10), respectively. These results are consistent with the close phylogenetic relationships between the three species, indicating this set of primers might proved useful for studying the levels of genetic diversity and population differentiation in these three species and in other phylogenetically close species of the genus Diplodus and Sparus.     

Characterization and transferability of microsatellite markers of the cultivated peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis>)

Background  

The genus Arachis includes Arachis hypogaea (cultivated peanut) and wild species that are used in peanut breeding or as forage. Molecular markers have been employed in several studies of this genus, but microsatellite markers have only been used in few investigations. Microsatellites are very informative and are useful to assess genetic variability, analyze mating systems and in genetic mapping. The objectives of this study were to develop A. hypogaea microsatellite loci and to evaluate the transferability of these markers to other Arachis species.     

Two highly validated SSR multiplexes (8‐plex) for Euphrates' poplar,Populus euphratica (Salicaceae)

Multiplex PCR amplification of microsatellites has significantly increased the throughput and decreased the costs of genotyping. We have developed two highly polymorphic microsatellite multiplexes for Populus euphratica, the only tree species found in desert regions of Western China and adjacent Central Asian countries. The first of these multiplex kits comprises an eight‐Plex of genomic SSRs (gSSRs) obtained from published databases. The second comprises an eight‐plex of newly designed EST‐SSRs (eSSRs) based on expressed sequence tags for P. euphratica. Both kits were tested on a sample of 170 individuals from four populations. The gSSRs exhibited slightly more polymorphism than the eSSRs. The new multiplex protocols yielded consistent results in the hands of multiple researchers, demonstrating their robustness. The 16 loci used in the kits exhibited a high transferability rate (82.0%) in eight other poplar species belonging to five different sections of the genus. Both kits should therefore be useful for further investigations of population genetics in P. euphratica and related species. Our results indicate that it is essential to follow recently established recommendations when developing microsatellite markers, including verifying the amplification efficiency, detecting null alleles and carefully measuring error rates.     

Isolation and characterization of polymorphic microsatellite loci from a repeat-enriched genomic library of stone flounder (Kareius bicoloratus) and cross-species amplification

Stone flounder (Kareius bicoloratus) is a rare fish species in China. Here, we reported 11 polymorphic microsatellite loci isolated from a repeat-enriched genomic library of stone flounder. The number of alleles, observed and expected heterozygosity per locus in 32 individuals ranged from 3 to 6, from 0.1613 to 0.8667 and from 0.1549 to 0.7932, respectively. Two 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 Kareius bicoloratus. Yong-Sheng Tian and Gui-Dong Miao are contributed equally.     

Isolation of microsatellite loci and cross‐species amplifications in three gobiid fish of the genus Pomatoschistus

Gobiids of the genus Pomatoschistus are increasingly being investigated as models for adaptation to coastal environments and for mating system studies. Among the dozen currently analysed species, microsatellite primers have been characterized only for Pomatoschistus minutus. This paper describes seven new polymorphic loci isolated from Pomatoschistus marmoratus and Pomatoschistus microps, two species that hybridize. Cross‐species amplification was tested for these new loci, together with seven already published P. minutus loci. Systematic amplification of samples of each of the three species provided a first indication of their polymorphism.     

Isolation and characterization of microsatellite loci in <Emphasis Type="Italic">Pedicularis verticillata</Emphasis> L. using PCR-based isolation of microsatellite arrays (PIMA)

Pedicularis verticillata L. is a highly valuable herb for traditional Chinese medical treatment. In this report, 11 microsatellite loci from P. verticillata were isolated. The simple sequence repeat (SSR) markers were screened in 23 samples of wild populations of P. verticillata, and eight samples from its sister P. ikomai. The number of alleles ranged from 4 to 13, and value of expected (H _E) and observed (H ₀) heterozygosity was 0.62609–0.89662 and 0–0.95652, respectively. All loci were significantly deviated from Hardy–Weinberg expectations due to the heterozygote deficiency, indicating a dramatic loss of genetic polymorphisms in the restrictedly distributed species. The markers amplified well in the two species are useful for examining genetic diversity and population genetic structure, which, in turn, can provide information for establishing conservation strategy for these endangered 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 11 polymorphic microsatellite loci from <Emphasis Type="Italic">Fatsia polycarpa</Emphasis> (Araliaceae), an element of evergreen forests in Taiwan

Fatsia polycarpa Hayata (Araliaceae) is an element of evergreen forests at middle elevations in Taiwan, also representing the southern limitation in distribution of the genus. In this study, we described the development of 11 microsatellite loci from F. polycarpa in Taiwan for genetic studies. These new markers were tested in 20 individuals of 5 populations of the woody species. The number of alleles ranged from 9 to 18. The expected (H _E ) and observed (H _O ) heterozygosities were 0.80–0.95 and 0.10–0.75, respectively. Six of the 11 microsatellite loci are significantly deviated from Hardy–Weinberg expectations likely due to the population structure within samples. No linkage disequilibrium were observed from pairwise comparisons of loci. Chi-Chun Huang, Kuo-Hsiang Hung, and Tsai-Wen Hsu contributed equally to this article.     

Development of 10 microsatellite loci for <Emphasis Type="Italic">Rheum tanguticum</Emphasis> (Polygonaceae)

Rhubarb is an important Traditional Chinese Medicine. However, the wild resource has been declining. In order to design appropriate conservation methods for the official species across their natural distributions, it is important to characterize their genetic diversity. Here, we describe the development of 10 new microsatellite loci for AC/TG/CCA in Rheum tanguticum. The microsatellites were enriched using the combined biotin capture method. The polymorphism of each locus was further assessed in 12 individuals from four geographically distinct populations of this species. The number of alleles ranged from three to seven and the expected heterozygosity ranged from 0.53 to 0.73. All markers have been checked in the other three species in the genus and two of them together comprise the official medicinal rhubarb resource with R. tanguticum. These microsatellite markers could provide a useful tool for genetic and conservation studies of the rhubarb species.     

Microsatellite multiplexes for high‐throughput genotyping of French grunts (Haemulon flavolineatum,Pisces: Haemulidae) and their utility in other grunt species

Grunts of the genus Haemulon are abundant in the western Atlantic and are an important component of regional fisheries. Management efforts for these and other Caribbean reef fish require an understanding of population structure and the extent of dispersal among populations. We characterized 11 polymorphic microsatellite loci for French grunts (Haemulon flavolineatum) and grouped them into three PCR multiplexes to facilitate high‐throughput genotyping. These loci will be useful for population genetic studies of French grunts. Cross‐species amplifications suggest these loci are also suitable for population genetic studies of H. aurolineatum, H. plumieri, and H. sciurus.     

Isolation and characterization of di‐ and tetranucleotide microsatellite loci in the yellow‐spotted night lizard Lepidophyma flavimaculatum (Squamata: Xantusiidae)

We report the development of 17 di‐ and tetranucleotide microsatellite markers in the Yellow‐Spotted Night Lizard Lepidophyma flavimaculatum. Levels of heterozygosity ranged from 0 to 100%. Several loci also amplify in other Lepidophyma species and several species of the sister genus Xantusia. High levels of variation at some loci indicate that they will be useful for parentage assessment and population genetic studies, whereas the 15 loci that amplify across multiple Lepidophyma species suggest these will be useful in determining the origin of parthenogenetic populations.     

Highly polymorphic microsatellite loci for the Parsley frog (Pelodytes punctatus): characterization and testing for cross-species amplification

A microsatellite library was developed using genomic DNA of the Parsley frog, Pelodytes punctatus, an amphibian species which inhabits Mediterranean temporary pond systems. Number of alleles and heterozygosity ranged from 10 to 25 and from 0.66 to 0.90, respectively. Cross-species amplification was successful for 13 of the 15 developed loci for the related species, Pelodytes ibericus. The high levels of polymorphism revealed by these loci will be extremely useful for characterizing the population genetic diversity and structure and to estimate levels of dispersal and gene flow in the species P. punctatus and P. ibericus.     

Microsatellite DNA loci from the typical halophyte <Emphasis Type="Italic">Thellungiella salsuginea</Emphasis> (Brassicaceae)

Thellungiella salsuginea (Brassiaceae) is a typical halophyte which can tolerate extreme cold, drought, and salinity. In order to understand the adaptive evolution of this species in the arid habitats, it is important to know its genetic structure. In this study, 17 polymorphic microsatellite loci were isolated and characterized from an enrichment genomic library of this species. We further assessed the polymorphisms of each locus in 18 individuals from nine geographically distant populations. The number of alleles per locus ranged from six to fourteen. The observed and expected heterozygosity ranged from 0.17 to 0.28 and 0.32 to 0.45, respectively. These markers have been crossly checked in another congeneric species, T. halophila. These microsatellite markers will be useful for investigating population genetics and adaptive evolution of this species and morphological divergence between and it and the closely related species.     

Isolation and characterization of polymorphic microsatellite markers in <Emphasis Type="Italic">Cupressus </Emphasis><Emphasis Type="Italic">chenggiana</Emphasis> S. Y. Hu (Cupressaceae)

Cupressus chenggiana S. Y. Hu (Cupressaceae) is an endemic and endangered conifer species in southwest China. In order to study the population genetics and design the effective conservation methods, we aimed to develop microsatellite primers for this species in the present study. We developed eight new microsatellite loci for this species through biotin capture method. Polymorphism of each locus was further assessed in 18 individuals from three geographically distant populations. The number of alleles per locus ranged from 6 to 11 with an average of 8.13. The observed and expected heterozygosities ranged from 0.219 to 0.296 and from 0.374 to 0.470, with averages of 0.254 and 0.417, respectively. We further found that three of nine microsatellite loci developed previously for another congeneric species showed polymorphic banding patters. We performed primer-crossing tests of these loci in the other two congeneric species which are closely related to C. chenggiana (C. gigantea and C. duclouxiana). These microsatellite markers would be effective for analyzing genetic diversity and population genetic structure of this species and its morphological differentiation with the close relatives.     

Phylogeography and genetic structure of the Canarian common chaffinch (Fringilla coelebs) inferred with mtDNA and microsatellite loci

The widespread common chaffinch (Fringilla coelebs) inhabits five of the seven Canary Islands. Sequences of the mitochondrial cytochrome b gene (1002 bp) revealed new insights into the systematics and phylogeography of this taxon. Additionally, a set of microsatellite loci were analyzed to examine the structure of these populations. Our results suggest that a new species of the genus Fringilla is present in the Canary Islands, which comprises at least three subspecies, but with a different distribution to that which has been morphologically accepted. The specimens from Gran Canaria are genetically distinct from those of La Gomera and Tenerife (F. c. canariensis), which suggests the existence of an undescribed taxon. Furthermore, nuclear microsatellite data suggest an ongoing incipient speciation process in this population. This study provides both important conservationist implications and a basis for re-evaluating the taxonomic status of the Canarian Fringilla coelebs populations.     

A multiplex panel of microsatellite markers for widespread sub‐Saharan rodents of the genus Mastomys

We isolated and characterized 12 polymorphic microsatellite loci in the sub‐Saharan rodent Mastomys huberti. We tested cross‐species amplification of all these loci in three closely related Mastomys species: M. coucha, M. erythroleucus and M. natalensis. Multiplex panels comprising 11 loci were developed and their application to a set of individuals in each species allowed clear and easy characterization of allele sizes. Statistics from 31 M. huberti coming from one locality in Mali showed no deviation from Hardy–Weinberg equilibrium except for one locus, and no significant linkage disequilibria between loci.     

Isolation and characterization of polymorphic microsatellite loci in Acanthoscelides obtectus Say (Coleoptera: Bruchidae)

Six microsatellite loci were isolated from the bruchid Acanthoscelides obtectus Say (Coleoptera: Bruchidae). Each locus was polymorphic, with the number of alleles ranging from 3 to 18. We found high levels of within‐population variation at most loci, with heterozygosities ranging from 0 to 0.75. Cross‐species amplification of these loci was tested in two other species of the genus Acanthoscelides, A. obvelatus Bridwell and A. argillaceus Sharp.