Development and characterization of microsatellite markers in black poplar (Populus nigra L.)

Using an enrichment procedure, we have cloned and sequenced microsatellite loci from black poplar (Populus nigra L.) and developed primers for sequence-tagged microsatellite (STMS) analysis. Twelve primer pairs for dinucleotide repeats produced fragments of sufficient quality which were polymorphic in P. nigra. Some of them also showed amplification in other Populus species (P. deltoides, P. tricocarpa, P. tremula, P. tremuloides, P. candicans, and/or P. lasiocarpa). The best nine and (GT) (GA) microsatellite markers were tested on a set of 23 P. nigra genotypes from all over Europe. The microsatellites were highly polymorphic, with 10–19 different alleles per microsatellite locus among these 23 genotypes. WPMS08 sometimes amplified three fragments. Using the other eight marker loci, the level of heterozygosity among the plants was on average 0.71 (range 0.25–1.00). The microsatellite markers developed will be useful for screening the genetic diversity in natural populations and in gene bank collections. Received: 21 October 1999 / Accepted: 24 November 1999     

Isolation and strategies of novel tetranucleotide microsatellites with polymorphisms from different chromosomes of the rhesus monkey (Macaca mulatta)

A total of 45 tetranucleotide chromosome-specific microsatellite markers with polymorphism were developed successfully based on three reference rhesus monkey genomes and on In-silico PCR prescreening. The polymorphic information content (PIC) values of 45 polymorphic microsatellite loci ranged from 0.487 to 0.879, with an average of 0.715, which were proven to be moderate to highly polymorphic. We detected 315 alleles on 45 microsatellite loci in 24 Rhesus monkeys. The number of alleles ranged from 3 to 15 and the mean number of alleles was 7 for each locus. Accordingly, the observed and expected heterozygosities obtained were between 0.417 and 1.0 and between 0.550 and 0.908, with an average value of 0.736 and 0.767, respectively. Genetic information demonstrated that 10 loci significantly deviated from Hardy–Weinberg equilibrium (P?<?0.05). All 45 primers were not significant with regard to linkage disequilibrium (P?>?0.001). Pearson correlation indicated that the PIC value exhibited a significant negative correlation with the loci number (r?=?? 0.741, P?=?0.022), whereas the positive correlation with the number of the samples (r?=?0.847, P?=?0.070) was not significant. This may be attributed to the presence of random particularities within the loci. The T test of the sample groups indicated that the PIC difference was not significant when the number of samples was set at 10 and/or?≥?15 (P?=?0.7472?~?0.8564). These polymorphic and valuable microsatellite loci will facilitate further conservation genetics studies for rhesus monkeys and can be further applied to develop novel genetic markers for other species.

     

Development of microsatellite markers for southern catfish (Silurus meriaionalis) and cross‐species amplification in northern sheatfish (Silurus soldatovi)

Forty microsatellite markers were developed from a (CA)_n enrichment library created from the DNA of southern catfish (Silurus meriaionalis). Also, the population structure of northern sheatfish (Silurus soldatovi) was examined by 24 microsatellite loci. They are polymorphic in at least one of the two geographically distant populations sampled from the Heilongjiang River and the Songhuajiang River in North China, respectively. Unbiased expected heterozygosity levels varied from 0.435 to 0.946 and number of alleles per locus varied from three to 20. Results indicated that these microsatellite loci were highly polymorphic and could be used as genetic markers.     

Novel tetranucleotide microsatellite markers from the Del Norte salamander (Plethodon elongatus) with application to its sister species the Siskiyou Mountain salamander (P. stormi)

Eleven tetranucleotide microsatellite loci were developed for the Del Norte salamander (Plethodon elongatus). The loci were variably polymorphic, ranging from two to 20 alleles per locus, with expected heterozygosities ranging from 0.07 to 0.86. The loci also amplified in a congener, the Siskiyou Mountain salamander (P. stormi). The microsatellite loci will be used to assess the utility of highly polymorphic markers to assay within‐ and between‐species differentiation between these two closely related species.     

Development of microsatellite markers for muskellunge (Esox masquinongy) and cross‐species amplification in two other esocids

We report the development of seven polymorphic microsatellite loci in muskellunge (Esox masquinongy) using an unenriched subgenomic library. Polymorphic loci exhibited 2–11 alleles with observed heterozygosities ranging from 0.190 to 0.917 (n = 24). All seven loci amplified by their respective primer pairs resulted in monomorphic products in northern pike (E. lucius) whereas three loci amplified monomorphic products in grass pickerel (E. americanus americanus); these results imply conservation of flanking sequence but loss of polymorphism between these closely related species. Only one of six microsatellite primers developed in a previous study in northern pike amplified polymorphic products in muskellunge.     

Isolation and characterization of microsatellite DNA markers for the flagfish <Emphasis Type="Italic">Jordanella </Emphasis><Emphasis Type="Italic">floridae</Emphasis>

The flagfish (Jordanella floridae) is commonly used in studies of wetlands ecology. Here we describe the isolation of ten microsatellite loci, six of which were polymorphic. The observed number of alleles ranged from 4 to 24 and observed heterozygosities ranged from 0.59 to 0.81 for the polymorphic loci. The isolation of these markers will enable estimations of genetic diversity in natural populations.     

Microsatellite loci for the southern pine beetle (Dendroctonus frontalis) and cross‐species amplification in Dendroctonus

We developed microsatellite loci for the southern pine beetle (Dendroctonus frontalis). Twelve microsatellite loci were identified. Eight loci were polymorphic and sufficiently variable in 62 individuals (expected heterozygosity ranged from 0.707 to 0.880) to investigate population structure. All loci conformed to HWE except Dfr‐14, which showed heterozygote excess, and no two loci deviated from linkage equilibrium. The loci were tested for cross‐species amplification in four species of Dendroctonus (D. valens, D. terebrans, D. brevicomis, and D. ponderosae). Seven loci were polymorphic in at least one of the species tested.     

Characterization of polymorphic microsatellite loci from Round Island Petrels (Pterodroma arminjoniana) and their utility in other seabird species

Six microsatellite loci were isolated from the petrels of Round Island, near Mauritius in the Indian Ocean (Pterodroma arminjoniana). Three loci were monomorphic in P. arminjoniana but were found to be polymorphic in other Procellariiforms. Cross-utility of all six loci was tested in 17 Procellariiform and 1 penguin species. In addition, 53 microsatellite loci developed for other species of birds were tested for cross-species amplification in P. arminjoniana. Six of these loci were found to be polymorphic.     

Development and characterization of microsatellite markers for parentage analyses of the coral reef damselfish (<Emphasis Type="Italic">Pomacentrus amboinensis</Emphasis>: Pomacentridae)

Five new polymorphic microsatellite loci were developed for the coral reef damselfish Pomacentrus amboinensis. Twenty-four individuals from two Great Barrier Reef populations were genotyped at the five loci, with numbers of alleles per locus ranging from 6–23 and observed heterozygosity between 0.42–0.92. In addition, the cross-species testing of six primers developed for Stegastes partitus revealed one primer (SpGATA40) that was also polymorphic for P. amboinensis. Due to high levels of polymorphism (≥14 alleles) in at least four of the six loci and a high proportion of tetranucleotide repeats, these microsatellite markers should be useful for parentage assignment as well as other investigations of individual relatedness.     

Isolation and characterization of microsatellites in yellow perch (Perca flavescens)

A total of 45 microsatellite loci from yellow perch, Perca flavescens, were isolated and characterized. Among the 45 microsatellite loci, 32 had more than two alleles. A wild population of P. flavescens (n = 48) was used to examine the allele range of the microsatellite loci. Mendelian inheritance of alleles was confirmed by examining the amplified products in pair‐mated families. The number of alleles for the 32 polymorphic loci varied from two to 16, and observed heterozygosity ranged between 0.024 (YP79) and 0.979 (YP60). Cross‐species polymorphic amplification in four other Percidae species was successful for 22 loci.     

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.     

Twenty‐seven new microsatellites for the migratory Asian catfish family Pangasiidae

In this paper, we describe primers for 27 new microsatellite loci tested on five species of migratory Asian catfish: Pangasius krempfi, P. bocourti, P. conchophilus, P. pleurotaenia, and Helicophagus waandersii. These primers were developed from a (GATA)_n library created from the pooled DNA of three species of pangasiid catfish. All 27 loci are polymorphic in at least one species. Fifteen loci are polymorphic in at least three species. The primers described in this paper are thus shown to be useful in several species within the catfish family Pangasiidae, and may prove useful in additional species in future tests.     

Genetic variation and population structure of endemic yellow catfish, <Emphasis Type="Italic">Horabagrus</Emphasis><Emphasis Type="Italic">brachysoma</Emphasis> (Bagridae) among three populations of Western Ghat region using RAPD and microsatellite markers

Random amplified polymorphic DNA (RAPD) and microsatellite markers were applied to evaluate the genetic variation in endemic and endangered yellow catfish, Horabagrus brachysoma sampled from three geographic locations of Western Ghat, South India river systems. In RAPD, of 32 10-mer RAPD primers screened initially, 10 were chosen and used in a comparative analysis of H. brachysoma collected from Meenachil, Chalakkudy and Nethravathi River systems. Of the 124 total RAPD fragments amplified, 49 (39.51%) were found to be shared by individuals of all 3 populations. The remaining 75 fragments were found to be polymorphic (60.48%). In microsatellites, six polymorphic microsatellite loci were identified by using primers developed for Pangasius hypophthalmus, Clarias macrocephalus and Clarias gariepinus. The identified loci were confirmed as microsatellite by sequencing after making a clone. The nucleotide sequences of 6 loci were published in NCBI genbank. The number of alleles across the six loci ranged from 4 to 7 and heterozygosities ranged from 0.07 to 0.93. The mean number of alleles and effective number of alleles per locus were 5.00 and 3.314, respectively. The average heterozygosity across all investigated samples was 0.72, indicating a significant deficiency of heterozygotes in this species. RAPD and microsatellite methods reported a high degree of gene diversity and genetic distances depicted by UPGMA dendrograms among the populations of H. brachysoma.     

Isolation and charaterization of polymorphic microsatellite loci from so-iuy mullet (Mugil soiuy Basilewsky 1855)

The first set of polymorphic microsatellite markers were developed from so-iuy mullet (Mugil soiuy Basilewsky 1855). From a (GT)n-enriched genomic library, 53 microsatellites were selected for designing microsatellite primers, of which 36 gave working primer pairs. Ten of these loci were polymorphic in a test population of 24 individuals with alleles ranging from 3 to 9, and observed and expected heterozygosities from 0.2083 to 0.9167 and from 0.2651 to 0.8812, respectively. No significant linkage disequilibrium between pairs of loci was found, but two loci significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction. These polymorphic microsatellite loci should provide sufficient level of genetic diversity to investigate the fine-scale population structure and evaluate the breeding strategy in Mullet. Genbo Xu and Changwei Shao Contributed equally.     

Isolation and characterization of microsatellites in Drosophila montana and their cross‐species amplification in D. virilis

We report the isolation of 20 microsatellite loci from Drosophila montana and their cross amplification in the relative D. virilis. All microsatellite loci were polymorphic in the focal species D. montana, with gene diversities ranging from 0.23 to 0.93. In D. virilis only eight loci (40%) amplified and two loci were polymorphic (10%). These markers represent the first report of microsatellites isolated in D. montana. They could be applied for studying population structure and phylogeography. The largest benefit, however, will be their use in studies of quantitative trait loci, such as the mapping of behavioural quantitative trait loci.     

Microsatellite DNA primers for the endangered vermilion darter,Etheostoma chermocki,and cross‐species amplification in other darters (Percidae: Etheostoma)

The endangered vermilion darter (Etheostoma chermocki) is endemic to the Black Warrior River system in the Mobile Basin in Alabama. Restoration and conservation of this species require an understanding of its population genetic structure, which can be characterized using microsatellite DNA. Nine microsatellite loci were developed; eight loci were polymorphic. Although observed heterozygosity was lower than expected heterozygosity in most polymorphic loci, only one locus showed significant deviation from Hardy–Weinberg equilibrium. These nine markers were tested in an additional 24 species of Etheostoma and appear to have sufficient allelic variation to be useful in studies of population genetic structure.     

Polymorphic microsatellite markers for the Iberian rock lizard species,Iberolacerta cyreni,and cross‐species priming in other Iberolacerta species

Eight polymorphic microsatellite loci are described for the Iberian rock lizard species, Iberolacerta cyreni. Loci were isolated from a partial genomic library that had been enriched for AAAG repeat sequence. Number of alleles per locus ranged from two to 13 in a sample of 24 individuals from the Sierra de Guadarrama (central Spain). Observed heterozygosity ranged from 0.13 to 0.96. At least three loci were amplified and polymorphic in four other Iberian rock lizard species: Iberolacerta monticola, Iberolacerta bonnali, Iberolacerta aranica and Iberolacerta aurelioi. These markers will be used to study mating strategies and reproductive success in I. cyreni.     

Genetic variation and phylogenetic relationship between two species of yellow catfish, <Emphasis Type="Italic">Horabagrus brachysoma</Emphasis> and <Emphasis Type="Italic">H. nigricollaris</Emphasis> (Teleostei: Horabagridae) based on RAPD and microsatellite markers

The two species of yellow catfish, Horabagrus brachysoma and H. nigricollaris are categorized as ‘endangered’ and ‘critically endangered’ respectively in their wild habitat. Proper knowledge of genetic structure and variability of these endangered species are highly essential for the management, conservation and improvement of fish stocks. Therefore, genetic variation and phylogenetic relationships between these species of yellow catfish sampled from Chalakkudy River in the hot spot of biodiversity-Western Ghats region, Kerala, India were analyzed by using Random amplified polymorphic DNA (RAPD) and microsatellite markers. 85 RAPD and five microsatellites loci were detected to analyze the genetic variation and phylogenetic relationships among these species. Out of 85 RAPD loci produced only 52.94% were polymorphic whereas in microsatellite, all 5 loci were polymorphic (100%). Species-specific RAPD bands were found in both species studied. In microsatellite, the number of alleles across the five loci ranged from 1 to 8. The observed heterozygosities in H. brachysoma and H. nigricollaris were 0.463 and 0.443, respectively. Here, both RAPD and microsatellite methods reported a low degree of gene diversity and lack of genetic heterogeneity in both species of Horabagrus which strongly emphasize the need of fishery management, conservation and rehabilitation of these species.     

Isolation and charaterization of 12 dinucleotide microsatellite loci from Belenger’s jewfish (Johnius belengerii Cuvier 1830)

We describe the first isolation of 12 polymorphic microsatellite markers from Belenger’s jewfish (Johnius belengnerii Cuvier 1830). From a (GT)n-enriched genomic library, 54 microsatellites were selected for designing microsatellite primers, of which 36 gave working primer pairs. 12 of these loci were polymorphic in a test population of 21 individuals with alleles ranging from 3 to 18, and expected and observed heterozygosities from 0.5772 to 0.9449 and from 0.4286 to 0.9231, respectively. No significant linkage disequilibrium between pairs of loci was found, however, loci Jobe24 significantly deviated from Hardy–Weinberg equilibrium after Bonferroni correction. These polymorphic microsatellite loci should provide sufficient level of genetic diversity to investigate population structure in Belenger’s jewfish.