Mining single nucleotide polymorphisms from EST data of hard clam Meretrix meretrix
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In this study, we describe the first set of single nucleotide polymorphism (SNP) genotyping assays for hard clam (Meretrix meretrix) that were discovered through expressed sequence tag (EST) database mining. A total of 3,129 ESTs were assembled and 228 putative SNPs were identified from 51 SNP-containing contigs, with an average of 0.64 SNPs per 100 bp. A/G (T/C) and A/C (T/G) types had a high frequency and accounted for 46.2% and 35.6%, respectively. Thirty-two SNP genotyping assays were designed for validation based on allele-specific polymerase chain reaction (PCR) and melting temperature (Tm)-shift analysis, and 15 (46.9%) of them showed polymorphisms with the minor allele frequency ranging from 0.075 to 0.475. Twelve (37.5%) assays failed due to amplification failure of one or both allele specific primers, and five (15.6%) assays lacked sufficient temperature discrimination for accurate genotype scoring. BLASTX gave significant hits for all 15 genotyped SNP-containing contigs but all were synonymous coding SNPs. These results indicate that EST data constitutes a valuable resource for SNP discovery and further maker development.
KeywordsMeretrix meretrix Single nucleotide polymorphism SNP AS-PCR Tm-shift
This research was supported by grants from Program of Liaoning Province Science and Technology Commission (No. 2008203002) and Program of Liaoning Ocean and Fisheries Department (No. 200801).
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