The amplicons were confirmed to be the alpha region of the MHC class I gene by comparing each determined sequence with the alpha region of the MHC class I sequences of fishes available in GenBank using the BLAST program (http://www.ncbi.nlm.nih.gov). The results showed that all of the amplicons were the alpha region of the MHC class I. Then they were aligned using the CLUSTAL W program, and gene diversity was estimated using MEGA version 4.0 (Tamura et al., 2007) and Arlequin version 3.01 (Excoffier et al., 2005).