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Using Next-Generation Sequencing and Cross-Species Amplification in the Genus Pseudopungtungia
National Institute of Biological Resources, Incheon 404-708, Korea
School of Life Science, Kyungpook National University, Daegu 702-701, Korea
Department of Biology, Teachers College & Institute for Phylogenomics and Evolution, Kyungpook National University, Daegu 702-701, Korea
* Author to whom correspondence should be addressed.
Received: 1 June 2013; in revised form: 10 July 2013 / Accepted: 18 September 2013 / Published: 1 October 2013
Abstract: Nuclear microsatellite markers for Pungtungia herzi were developed using a combination of next-generation sequencing and Sanger sequencing. One hundred primer sets in the flanking region of dinucleotide and trinucleotide repeat motifs were designed and tested for efficiency in polymerase chain reaction amplification. Of these primer sets, 16 new markers (16%) were successfully amplified with unambiguous polymorphic alleles in 16 individuals of Pungtungia herzi. Cross-species amplification with these markers was then examined in two related species, Pseudopungtungia nigra and Pseudopungtungia tenuicorpa. Fifteen and 11 primer pairs resulted in successful amplification in Pseudopungtungia nigra and Pseudopungtungia tenuicorpa, respectively, with various polymorphisms, ranging from one allele (monomorphic) to 11 alleles per marker. These results indicated that developing microsatellite markers for cross-amplification from a species that is abundant and phylogenetically close to the species of interest is a good alternative when tissue samples of an endangered species are insufficient to develop microsatellites.
Keywords: microsatellite; Pungtungia herzi; Pseudopungtungia; cross-species amplification; next-generation sequencing
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Yun, Y.-E.; Yu, J.-N.; Kim, S.K.; Hwang, U.W.; Kwak, M. Using Next-Generation Sequencing and Cross-Species Amplification in the Genus Pseudopungtungia. Int. J. Mol. Sci. 2013, 14, 19923-19931.
Yun Y-E, Yu J-N, Kim SK, Hwang UW, Kwak M. Using Next-Generation Sequencing and Cross-Species Amplification in the Genus Pseudopungtungia. International Journal of Molecular Sciences. 2013; 14(10):19923-19931.
Yun, Young-Eun; Yu, Jeong-Nam; Kim, Sang K.; Hwang, Ui W.; Kwak, Myounghai. 2013. "Using Next-Generation Sequencing and Cross-Species Amplification in the Genus Pseudopungtungia." Int. J. Mol. Sci. 14, no. 10: 19923-19931.