Characterization of fourteen single nucleotide polymorphism markers in the Manila clam ( Ruditapes philippinarum )
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TECHNICAL NOTE
Characterization of fourteen single nucleotide polymorphism markers in the Manila clam (Ruditapes philippinarum) Hongtao Nie • Lianhui Liu • Jianfeng Ding Xiwu Yan
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Received: 23 July 2014 / Accepted: 29 July 2014 / Published online: 9 August 2014 Ó Springer Science+Business Media Dordrecht 2014
Abstract Fourteen single nucleotide polymorphisms (SNPs) were developed and characterized from expressed sequence tag sequence of the Manila clam, Ruditapes philippinarum. All loci had two alleles and the minor allele frequency ranged from 0.146 to 0.467. The observed and expected heterozygosities ranged from 0.259 to 0.792 and from 0.252 to 0.503. These are the first SNPs developed for assessing genetic variation and population structure in the Manila clam and will provide a useful complement to currently available genetic markers. Keywords Ruditapes philippinarum Manila clam SNP Tetra-primer ARMS-PCR The Manila clam (Ruditapes philippinarum), naturally occurring in Japan, China, and Korea, is now a worldwide species of clam cultivation industries. However, the wild stocks of R. philippinarum have been declining dramatically for last decades due to over-exploitation and the deterioration of environmental conditions in China. In recent years, recovery efforts such as artificial breeding program and stock enhancement are in progress (Zhang and Yan 2009). But, the genetic effects of hatchery individuals on wild populations of R. philippinarum have not yet been fully evaluated. Therefore, reasonable stock
H. Nie L. Liu J. Ding X. Yan (&) Engineering and Technology Research Center of Shellfish Breeding in Liaoning Province, Dalian Ocean University, Dalian 116023, China e-mail: [email protected] H. Nie Key Laboratory of Mariculture and Stock Enhancement in North China’s Sea, Ministry of Agriculture, Dalian Ocean University, Dalian 116023, China
management and genetic improvement are required for sustainable development of the clam aquaculture industry. To date, genetic markers for population studies of R. philippinarum have been generally limited to allozymes (Vargas et al. 2008), and microsatellites (An et al. 2012; Mura et al. 2012). In view of potential limitations of these approaches, we sought to add an additional class of markers, single nucleotide polymorphisms (SNPs), which possess a number of advantages for the study of population structure (Morin et al. 2004). Here, we report the first set of SNP markers in R. philippinarum which will provide a useful complement to currently available genetic markers. Polymorphic sites were identified from expressed sequence tag (EST) of R. philippinarum. A total of 6,468 EST sequences of R. philippinarum were downloaded from GenBank (Jan 6, 2013). The EST dataset was aligned and assembled using SeqMan Pro sequence assembly software (DNASTAR Inc., Madison, WI, USA). The contigs that contained four or more sequences were identified for searching candidate SNPs upon visual inspection. A singlebase mutation that occurred in two or more ESTs and was surroun
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