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Int. J. Mol. Sci. 2013, 14(10), 19923-19931; doi:10.3390/ijms141019923
Article

Using Next-Generation Sequencing and Cross-Species Amplification in the Genus Pseudopungtungia

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Received: 1 June 2013 / Revised: 10 July 2013 / Accepted: 18 September 2013 / Published: 1 October 2013
(This article belongs to the Section Molecular Pathology)
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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 microsatellite; Pungtungia herzi; Pseudopungtungia; cross-species amplification; next-generation sequencing
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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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.

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