Polymorphic microsatellite DNA markers in the Asiatic black bear <i>Ursus thibetanus</i>

Kitahara, Eiji; Isagi, Yuji; Ishibashi, Yasuyuki; Saitoh, Takashi

doi:10.1046/j.1365-294x.2000.01030.x

Cited by 90 publications

(32 citation statements)

References 11 publications

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“…Genotypes at 10 microsatellite DNA loci (G1A, G1D, G10B, G10L, G10M, G10X, MSUT-1, MSUT-2, MSUT-6, and MSUT-7;Paetkau et al 1995;Kitahara et al 2000) were determined for all individuals by the PCR technique. PCR amplification was carried out by using DNA thermal cyclers (Perkin Elmer 9700 and MJ Research, Inc. PTC-100) with 10 ll of reaction mixture containing 50 mM KCl, 1.5 mM MgCl 2 , 10 mM Tris-HCl (pH 8.3), 0.2 mM dNTP, 0.5 lM of each primer, and 0.5 units Taq DNA polymerase (Applied Biosystems AmpliTaq Gold).…”

Section: Dna Extraction and Genetic Typingmentioning

confidence: 99%

“…After denaturation of the sample at 95°C for 10 min, PCR amplification was performed for 30 cycles under the following conditions: 30 s at 95°C, 30 s at the locusspecific annealing temperature, and 30 s at 72°C. The locus-specific annealing temperatures were 65°C for G1A and G10B, 58°C for G1D and G10L, 54°C for G10M and G10X, and according to Kitahara et al (2000) for MSUT-1, -2, -6, and -7. PCR products were analyzed with a PRISM 377 or 3100-Avant Genetic Analyzer (Applied Biosystems), and genotypes were determined with GENOTYPER 2.1 or GENEMAPPER 3.5 software (Applied Biosystems).…”

Section: Dna Extraction and Genetic Typingmentioning

confidence: 99%

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Low genetic diversities in isolated populations of the Asian black bear (Ursus thibetanus) in Japan, in comparison with large stable populations

et al. 2007

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Populations of the Asian black bear (Ursus thibetanus) are relatively large and continuous in central Honshu, the main island of Japan, but they are isolated in western Honshu. To clarify the degree of genetic isolation of the populations in western Honshu, we compared the genetic diversities of four populations in western Honshu with that of one of the continuous populations of central Honshu. Three of the four western Honshu populations were isolated and the other was continuous with the central Honshu populations on a geographical distribution basis. The genotypes at 10 microsatellite loci of the sampled individuals were determined and the genetic structures of the populations examined. Genetic diversities were significantly lower in the isolated populations than in the continuous populations. The continuous population in central Honshu had high levels of genetic diversity, comparable to those in populations of the American black bear (Ursus americanus) and the brown bear (Ursus arctos). The genetic distances between the two continuous populations were smallest, even though their geographic distance was largest (>200 km) among all the pairs of neighboring populations examined. Low genetic diversity within the isolated populations suggested genetic drift due to the small population size; the genetic differentiation among the populations indicated low rates of gene flow among them.

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Section: Dna Extraction and Genetic Typingmentioning

confidence: 99%

Section: Dna Extraction and Genetic Typingmentioning

confidence: 99%

Low genetic diversities in isolated populations of the Asian black bear (Ursus thibetanus) in Japan, in comparison with large stable populations

et al. 2007

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“…We screened 30 autosomal microsatellite loci that had originally been developed for brown bears (Paetkau, Shields & Strobeck, 1998;Kleven et al, 2012), American black bears Paetkau & Strobeck, 1994), Asiatic black bears (Kitahara et al, 2000) or polar bears (Paetkau et al, 1995) for utility and variability in 58 polar bear samples covering subpopulations from all four ecoregions (Amstrup et al, 2008). Seven loci failed in PCR or were monomorphic in polar bears, leaving 23 microsatellite loci (Tables S2 and S3) for all following analyses.…”

Section: Autosomal Microsatellitesmentioning

confidence: 99%

High genetic variability of vagrant polar bears illustrates importance of population connectivity in fragmented sea ice habitats

et al. 2016

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Projections by the Intergovernmental Panel on Climate Change (IPCC) and sea ice forecasts suggest that Arctic sea ice will decline markedly in coming decades. Expected effects on the entire ecosystem include a contraction of suitable polar bear habitat into one or few refugia. Such large-scale habitat decline and fragmentation could lead to reduced genetic diversity. Here we compare genetic variability of four vagrant polar bears that reached Iceland with that in recognized subpopulations from across the range, examining 23 autosomal microsatellites, mitochondrial control region sequences and Y-chromosomal markers. The vagrants' genotypes grouped with different genetic clusters and showed similar genetic variability at autosomal microsatellites (expected heterozygosity, allelic richness, and individual heterozygosity) as individuals in recognized subpopulations. Each vagrant carried a different mitochondrial haplotype. A likely route for polar bears to reach Iceland is via Fram Strait, a major gateway for the physical exportation of sea ice from the Arctic basin. Vagrant polar bears on Iceland likely originated from more than one recognized subpopulation, and may have been caught in sea ice export during long-distance movements to the East Greenland area. Although their potentially diverse geographic origins might suggest that these vagrants encompass much higher genetic variability than vagrants or dispersers in other regions, the four Icelandic vagrants encompassed similar genetic variability as any four randomly picked individuals from a single subpopulation or from the entire sample. We suggest that this is a consequence of the low overall genetic variability and weak range-wide genetic structuring of polar bears -few dispersers can represent a large portion of the species' gene pool. As predicted by theory and our demographic simulations, continued gene flow will be necessary to counteract loss of genetic variability in increasingly fragmented Arctic habitats. Similar considerations will be important in the management of other taxa that utilize sea ice habitats.

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“…With microsatellites that isolated from Japanese black bear (U. thibetanus japonicus) (Kitahara et al 2000) and other Ursids (Paetkau et al 1995;Taberlet et al 1997), the tetranucleotide microsatellites we isolated should provide an ideal genetic tool kit to study the population genetics of the endangered Formosan black bear and other Asiatic black bears that are also under threat.…”

Section: Locusmentioning

confidence: 99%

Ten novel tetranucleotide microsatellite DNA markers from Asiatic black bear, Ursus thibetanus

Shih

Huang

et al. 2009

Conserv Genet

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Ten polymorphic microsatellite markers were developed for the endangered Formosan black bear (Ursus thibetanus formosanus) from a partial genomic library enriched for GAAA repeat. Polymorphism of these loci was evaluated in 27 Formosan black bear specimens of unknown relationship. The number of alleles per locus ranged from 5 to 15 and the observed heterozygosity of each locus ranged from 0.556 to 0.889. These loci should provide useful molecular tools to study conservation genetics of the Formosan black bear and other Asiatic black bears.Keywords Tetranucleotide microsatellite Á Tailed primers Á Ursus thibetanus Á Formosan black bear The Formosan black bear (Ursus thibetanus formosanus) is an endemic subspecies of Asiatic black bear inhabiting Taiwan (Wozencraft 2005). Similar to all other Asiatic black bears, degradation and fragmentation of habitat as well as poaching have caused a decrease in population and distribution of the Formosan black bear (Wang 1999;Hwang and Wang 2006). To formulate proper conservation strategies, it is important to understand the genetic diversity and genetic structure within and among populations of this subspecies.In this study, we reported ten novel easy-scored polymorphic tetranucleotide repeat (GAAA) microsatellite loci from the Formosan black bear. We followed the protocol developed by Hsu et al. (2003) to enrich microsatellitecontained fragment in a partial genomic library. The library was constructed from genomic DNA which extracted from tissue sample of a Formosan black bear individual using the proteinase K-chloroform method (Sambrook et al. 1989). Microsatellite-enriched PCR (polymerase chain reaction) library was ligated into pGEM-T Easy vector (Promega) and transformed into Escherichia coli DH5a. A total of 880 clones were lifted to Hybond-N ? membranes (Amersham Pharmacia Biotech) and hybridized with [c 32 P] ATP end-labelled (GAAA) 10 oligonucleotides, then 56 hybridized clones were sequenced using DYEnamic ET Dye Terminator Cycle Sequencing Kit for MegaBACE (Amersham Bioscience) on a MegaBACE 1000 autosequencer (Amersham Bioscience). Sequences were proofread using software SEQUENCER 4.2 (Gene Codes). We found 47 clones with microsatellite motif, of which 33 loci containing more than 10 units of GAAA motif were chosen to design the PCR primers.All forward primers were 5 0 -tailed with an M13-tail (5 0 -GGAAACAGCTATGACCAT-3 0 ) or a CAG-tag (5 0 -CAG TCGGGCGTCATCA-3 0 ) (Schuelke 2000; Boutin-Ganache et al. 2001). DNA extracted from tissue samples of 17 Formosan black bears and from faecal samples of ten Formosan black bears with unknown relationship were used to characterize these 33 loci. PCRs were set up in

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Polymorphic microsatellite DNA markers in the Asiatic black bear Ursus thibetanus

Cited by 90 publications

References 11 publications

Low genetic diversities in isolated populations of the Asian black bear (Ursus thibetanus) in Japan, in comparison with large stable populations

Low genetic diversities in isolated populations of the Asian black bear (Ursus thibetanus) in Japan, in comparison with large stable populations

High genetic variability of vagrant polar bears illustrates importance of population connectivity in fragmented sea ice habitats

Ten novel tetranucleotide microsatellite DNA markers from Asiatic black bear, Ursus thibetanus

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