Isolation and characterization of microsatellite loci in the Pacific pleasure oyster,Crassostrea corteziensis,and their cross‐species amplification in four other oyster species

Microsatellite loci were isolated in Crassostrea corteziensis using (GT)_n, (CT)_n and (CTGT)_n‐enriched genomic libraries. Within each of 45 sequenced clones, an average of three microsatellite regions (156 total) were observed. Thirty‐three primers were designed, from which 11 microsatellite loci amplified. Ten of those were polymorphic, with a range of two to 30 alleles. Three loci were not in Hardy–Weinberg equilibrium, and linkage disequilibrium was found for six pairs of loci. These microsatellite loci will be further tested for segregation distortions and null alleles to establish a set for population genetic studies of the species in the Northwest coasts of Mexico, and for optimization of aquaculture development. Seven of the microsatellite loci cross‐amplified in Crassostrea palmula, a sympatric species, and will be useful in further genetic studies.     

Characterization of microsatellite loci in the Jamaican fruit‐eating bat Artibeus jamaicensis and cross‐species amplification

Artibeus jamaicensis is one of the most common bat species in the neotropics, with a well‐defined polygynous social structure in caves. In order to study behaviour and to examine patterns of paternity and relatedness between different harem groups, we developed 14 microsatellite loci from two different enriched genomic libraries. We screened 125 individuals from two different bat colonies and found that polymorphism ranged from five to 13 alleles. Heterozygosity ranged from 63 to 95%. The primers amplified across 14 bat species, indicating their potential utility for population‐level studies in several closely related bat species.     

Development of microsatellite markers for the endangered semi‐shrub Chimaphila umbellata (Ericaceae)

Based on the DNA sequencing reads obtained using 454 pyrosequencing, primers amplifying 16 microsatellite loci were developed for the endangered semi‐shrub Chimaphila umbellata, which occurs sporadically in the Japanese Archipelago. These 16 loci were polymorphic in the populations sampled from the Hokkaido and Tohoku Districts; the mean number of alleles was 3.31 and 3.44, and the mean expected heterozygosity was 0.42 and 0.44, respectively. These loci were not linked to each other and contained no null alleles. Amplification using these primers was also tested in the congeneric species C. japonica, but only three of them successfully amplified DNA of the species. These markers will be used to examine genetic diversity and genetic differentiation in populations of C. umbellata.     

Microsatellite markers isolated from polyploid wood‐sorrel Oxalis alpina (Oxalidaceae)

Twelve polymorphic microsatellite loci were isolated from polyploid alpine wood‐sorrel, Oxalis alpina (Oxalidaceae), and optimized for future studies of its breeding system. The loci were screened for variability among 72 individuals from Pinos Altos, New Mexico. The primers amplified loci with allele number ranging from two to 17 per locus and with estimates of Nei's genetic diversity varying from 0.10 to 0.99. These primers provide an opportunity to use polymorphic DNA markers to study the causes of breeding system variability in this species.     

Primers from the orders Osteoglossiform and Siluriform detect polymorphic microsatellite loci in sun-catfish, Horabagrus brachysoma (Teleostei: Bagridae)

Horabagrus brachysoma (sun‐catfish, Bagridae, Siluriformes) is a valuable ornamental and food fish. The stock structure of H. brachysoma, necessary to conserve its declining natural populations, is not known. Twenty‐five primers developed for four fish species belonging to the orders Siluriform (3) and Osteoglossiform (1) were tested and eight primers amplified microsatellite loci in H. brachysoma. The results demonstrate that cross‐priming between fish species belonging to different families and even to different orders can yield microsatellite loci. Five of eight primers each amplified two loci. However, the loci that had repeat motifs after sequencing were considered only for genotyping. Finally, eight loci were polymorphic with hree to seven alleles. Individual fish genotype data (n = 42; 21 each in two rivers) at each locus was analysed. Significant genetic heterogeneity was detected at six loci. The identified loci exhibited potential for use in population genetics application in H. brachysoma.     

Development of sequence‐tagged microsatellite site markers for chickpea (Cicer arietinum L.)

Microsatellite loci were identified from chickpea (Cicer arietinum L.), the third most important grain legume crop in the world. A total of 13 sequence‐tagged microsatellite markers were developed using two different approaches: (i) amplification using degenerate primers and (ii) cloning of intersimple sequence repeat (ISSR)‐amplified fragments. Thirty‐five chickpea accessions were analysed, which resulted in a total of 30 alleles at the 13 loci. The observed heterozygosity ranged from 0.1143 to 0.4571 with an average of 0.2284. The cross‐species transferability of the sequence‐tagged microsatellite site (STMS) markers was checked in Cicer reticulatum, the wild annual progenitor of chickpea. These microsatellite markers will be useful for assessing the genetic diversity patterns within chickpea as well as aid in construction of intra‐ and interspecific genetic linkage maps.     

Microsatellite markers for testing host plant‐related population differentiation in the moth genus Yponomeuta

Nine microsatellite primers were developed for Yponomeuta padellus (Lepidoptera: Yponomeutidae) and tested for their applicability in analysing genetic population structure. Eight of the nine loci were highly polymorphic with on average 11.4 alleles. Cross‐species amplification of the nine primer pairs was tested in five other moth species. Primer pairs amplified in Y. cagnagellus, Y. malinellus, Y. evonymellus, and Y. rorellus but not in Y. sedellus and Plutella xylostella.     

Isolation and characterization of microsatellite loci in Penstemon rostriflorus (Plantaginaceae) and cross‐species amplification

Eight novel microsatellite primer pairs are presented for Penstemon rostriflorus, representing the first microsatellite markers available for this genus. Loci were characterized for 20 individuals from two populations in the Great Basin, USA. All loci are polymorphic within P. rostriflorus (seven to 13 alleles per locus; observed heterozygosity between 0.40 and 0.95), and therefore useful for population genetic studies within the species. Cross‐species transferability was tested on 40 additional species of Penstemon, and results indicate that these primers pairs will likely be useful for population genetic studies on many Penstemon species.     

Microsatellite loci for studies on population differentiation and connectivity of the red‐bellied yellow tail fusilier,Caesio cuning (Caesionidae)

Ten polymorphic microsatellites were isolated from the red‐bellied yellow tail fusilier Caesio cuning, a reef‐associated fish which occurs widely in the Indo‐Pacific region. The species is exploited by both small‐scale and commercial fisheries. Fifty individuals from six populations were genotyped using primers that reliably amplified 10 polymorphic loci. The number of alleles per locus ranged from three to 11. Observed heterozygosity (H_O) and expected heterozygosity (H_E) ranged between 0.400 to 0.880 and 0.337 to 0.843, respectively. These microsatellite loci may be used to study population structure, genetic diversity and connectivity of C. cuning in the range of its distribution.     

Microsatellite primers for fungus‐growing ants

We isolated five polymorphic microsatellite loci from a library of two thousand recombinant clones of two fungus‐growing ant species, Cyphomyrmex longiscapus and Trachymyrmex cf. zeteki. Amplification and heterozygosity were tested in five species of higher attine ants using both the newly developed primers and earlier published primers that were developed for fungus‐growing ants. A total of 20 variable microsatellite loci, developed for six different species of fungus‐growing ants, are now available for studying the population genetics and colony kin‐structure of these ants.     

Development of microsatellite markers for the red tide‐forming harmful species Chattonella antiqua,C. marina,and C. ovata (Raphidophyceae)

We have developed 11 microsatellite markers that are specific to Chattonella antiqua, C. marina, and C. ovata, the red tide‐forming harmful phytoplanktons. The 11 loci were amplified in the three species. The number of alleles per locus ranged from 5 to 16. The three species shared most microsatellite regions, although the genetic differences in specific loci were detected among them. These markers of the Chattonella species will be beneficial for biogeographical, detailed taxonomic, studies.     

Isolation and characterization of microsatellites in Neogobius kessleri (Perciformes,Gobiidae) and cross‐species amplification within the family Gobiidae

Fourteen polymorphic microsatellites were isolated from Neogobius kessleri, a benthic fish of Ponto‐Caspian origin which has been recently introduced into the Middle and Upper Danube River. Number of alleles and heterozygosity per locus in a sample of 32 fish individuals ranged from two to four and from 0.13 to 0.75, respectively. These primers will be useful in determining the population structure of N. kessleri. In addition, successful cross‐amplification was obtained for four related species, N. melanostomus, N. fluviatilis, N. gymnotrachelus and Proterorhinus marmoratus. These microsatellite loci may be useful for the evaluation of the origin of non‐native goby populations.     

Identification of polymorphic microsatellite markers from RAPD product in turbot (Scophthalmus maximus) and a test of cross‐species amplification

Five polymorphic microsatellite loci have been isolated and characterized from random amplified polymorphic DNA product in turbot, Scophthalmus maximus. Twelve microsatellites were selected for designing microsatellite primers, of which five gave working primer pairs. They had between four and nine alleles. Observed and expected heterozygosities varied from 0.76 to 0.90 and from 0.63 to 0.83, respectively. Five additional fish species assessed for cross‐species amplification revealed between one and three positive amplifications and between zero and two polymorphic loci per species.     

Isolation and characterization of microsatellite loci from the orchid Serapias vomeracea (Orchidaceae) and cross‐priming to other Serapias species

In this paper we describe the isolation and characterization of six polymorphic microsatellite loci from the orchid Serapias vomeracea. This species is widely distributed in the Mediterranean region. Microsatellite loci were isolated from an enriched library and primer pairs were designed for 18 loci. Primer pairs for six loci amplified well and were tested on samples from southern Italy. Levels of genetic variability detected at these six loci are high, with numbers of alleles per locus ranging from 3 to 6, and observed heterozygosity (H_O) ranging from 0.35 to 0.86. All primer pairs tested amplified DNA from four other Serapias species, indicating that the primers are useful for population genetic studies throughout the genus.     

Characterization of 35 microsatellite loci in the Pacific lion‐paw scallop (Nodipecten subnodosus) and their cross‐species amplification in four other scallops of the Pectinidae family

Four microsatellite‐enriched DNA libraries yielded 35 microsatellite loci from 100 primer pairs designed for Pacific lion‐paw scallop, Nodipecten subnodosus. The number of alleles ranged from four to 28. Three of the 35 loci were not in Hardy–Weinberg equilibrium and linkage disequilibrium was found for one pair of loci. These microsatellites will be used to analyse the population structure of the species in Mexico's Baja Peninsula to propose management strategies for scallop aquaculture development. Twenty‐six primer pairs cross‐amplified in Nodipecten nodosus, whereas none (Argopecten ventricosus) or few cross‐amplified in the Argopecten species.     

Isolation and characterization of Ligustrum micranthum (Oleaceae) microsatellite loci using paired‐end Illumina reads

Twenty‐six microsatellite loci were developed and characterized for Ligustrum micranthum, a species endemic to the Ogasawara Islands, Japan. The genetic structure of this species must be clarified in order to restore the island's ecosystem. A total of 8511 primer pairs were designed from de novo sequencing. Of the 48 primer pairs selected, amplification and polymorphisms were tested using one population each from the Chichijima and Hahajima Islands of the Ogasawara Islands. Twenty‐six microsatellite loci were successfully amplified and the number of alleles for these loci ranged from five to 31 per locus, and the mean expected heterozygosities were 0.858 and 0.849, respectively. No significant deviation from the Hardy–Weinberg equilibrium was observed in either population, and no significant linkage disequilibrium was detected between any locus pair. The microsatellite loci reported in this study can be used in future studies to evaluate the genetic structure and mating system of L. micranthum.     

Isolation and characterization of ten microsatellite loci in stingless bee Trigona spinipes (Apidae: Meliponini)

Stingless bees are the most abundant pollinators of Brazilian tropical flora. Trigona spinipes has some of the largest colonies of any stingless bee species found in several types of environment. This work describes the isolation and characterization of microsatellite loci for this species. A microsatellite‐enriched genomic library was constructed and ten primer pairs were designed for T. spinipes. The primers were tested in 20 unrelated individuals. The mean number of alleles was 8.10 and mean observed and expected heterozygosity were 0.655 and 0.680, respectively. Primers were also tested in cross‐species amplification and five loci were successfully amplified in Trigona chanchamayoensis, Trigona hyalinata, Tetragonisca angustula, Partamona mulata and Frieseomelitta varia. The microsatellite primers described herein will be useful for evaluating genetic variability and gaining a better understanding of the population structure of T. spinipes as well as other species of stingless bees.     

Cave insects with sex‐reversed genitalia had their most recent common ancestor in West Gondwana (Psocodea: Prionoglarididae: Speleketorinae)

The divergence date and ancestral distributional area of the psocid subfamily Speleketorinae, which includes taxa with reversed genitalia (female penis and male vagina of Afrotrogla and Neotrogla, tribe Sensitibillini), were estimated. The most basal divergence of the subfamily (between the North American Speleketor and the tribe Sensitibillini) was estimated to have occurred according to the separation between the North American continent and Gondwana, ca. 175 Ma. The most basal divergence of Sensitibillini (between African Afrotrogla + Sensitibilla and Brazilian Neotrogla) was estimated to have occurred according to the split of West Gondwana (separation between the African and South American continents), ca. 127 Ma. The biome of the ancestral distributional area of Sensitibillini (inland of West Gondwana) is believed to be arid to semi‐arid, which might strengthen the reversed sexual selection and then facilitate the origin of preadaptive features related to the evolution of a female penis. All extant Sensitibillini species inhabit carbonatic caves, but geological evidence suggested independent shifts of these genera to the carbonatic cave habitat in the Tertiary/Quaternary.     

Novel polymorphic microsatellite markers for paternity analysis in the red‐capped robin (Petroica goodenovii: Aves)

Seven microsatellite loci were isolated and characterized from the red‐capped robin Petroica goodenovii, using nonradioactive polymerase chain reaction (PCR)‐based techniques to screen an enriched genomic library. Five loci showed no evidence of null alleles and were variable [mean heterozygosity (H_E) = 0.440, mean number of alleles = 8]. Cross‐amplification using primers for microsatellites in Phylloscopus occipitalis and Emberiza schoeniclus yielded another two polymorphic loci. The combined set of five red‐capped robin and two cross‐amplified loci are suitable for paternity assignment (exclusion probability for seven unlinked loci = 0.9760).     

Cross-species amplification of 44 microsatellite loci developed for Chaetodon trifascialis, C. lunulatus and C. vagabundus in 22 related butterflyfish species

Forty‐four microsatellite primers developed for three species of butterflyfish were cross‐tested against 22 related confamilial species. Amplification success and cross‐species transferability of these markers were moderately high. Between 24 and 37 loci were amplified successfully in each species, with a mean success rate per species of 71.7% (± 1.8 SE). Rates of amplification success were comparable among primers designed for the three source species, ranging from a mean success rate per species of 16.9 loci (± 0.8 SE) for Chaetodon trifascialis source loci to 13.7 loci (± 1.5 SE) for C. vagabundus source loci. Polymorphism rates were high (76.1%± 3.1 SE of all successfully amplified loci), and 10 loci were polymorphic in all successfully amplified species (Tri14, B11, C5, D3, D113, D6, D117, D120, D111, D118). The number of alleles per polymorphic locus ranged from 2 to 8, and the average number of alleles across all polymorphic loci and all species was 3.6 (± 0.07 SE). Polymorphism rates were higher overall in primers designed for C. vagabundus (89.9%± 3.9 SE). Overall cross‐testing success was lowest for Heniochus chrysostomus, the most phylogenetically divergent species. The significant cross‐testing reported here provides a valuable resource that will enable population genetics studies to be undertaken on a range of butterflyfishes without the need for expensive and time‐consuming de novo microsatellite development.