Identification of Genes Involved in Immune Response, Microsatellite, and SNP Markers from Expressed Sequence Tags Generated from Hemocytes of Freshwater Pearl Mussel (Hyriopsis cumingii)

Abstract

Triangle sail mussel (Hyriopsis cumingii) is the most important mussel species commercially exploited for freshwater pearl production in China. However, its genome research is still at the infantry. Genomic resources for this species are largely not available. The objectives of this study was to generate expressed sequence tags from a hemocyte cDNA library, to identify genes involved in defense mechanisms, and to identify polymorphic markers from the expressed sequence tag (EST) resources for genetic analysis. A total of 5,290 ESTs were sequenced, obtaining 481 contigs and 1,165 singletons. BLAST similarity analysis indicated almost half (46.5%) of these ESTs were homologs of known genes while 53.5% were transcripts of unknown identities. Based on sequence similarities, 50 genes were identified as putative genes involved in immune and defense functions such as hemocyte immune process, stress proteins, adhesive proteins, proteases and protease regulators, antimicrobial peptides, lysosomal enzymes, cell apoptosis, and cell cycle proteins. A total of 201 microsatellites were identified from these ESTs, with 31 having sufficient flanking sequences for primer design. Polymerase chain reaction amplification was successful for 18 primer pairs and 14 of them were polymorphic. A total of 987 putative single nucleotide polymorphisms were identified including 204 transitions, 611 transversions, and 172 indels; 12 of them were involved in nine genes of defense mechanisms. These resources provide the material basis for future marker validation and genetic linkage and quantitative trait loci analysis in the freshwater pearl mussel.