Locus number estimation of MHC class II B in stone flounder and Japanese flounder

Members of major histocompatibility complex (MHC) family are important in immune systems. Great efforts have been made to reveal their complicated gene structures. But many existing studies focus on partial sequences of MHC genes. In this study, by gene cloning and sequencing, we identified cDNA sequences and DNA sequences of the MHC class II B in two flatfishes, stone flounder (Kareius bicoloratus) and homozygous diploid Japanese flounder (Paralichthys olivaceus). Eleven cDNA sequences were acquired from eight stone flounder individuals, and most of the polymorphic sites distributed in exons 2 and 3. Twenty-eight alleles were identified from the DNA fragments in these eight individuals. It could be deduced from their Bayesian inference phylogenetic tree that at least four loci of MHC class II B exist in stone flounder. The detailed whole-length DNA sequences in one individual were analyzed, revealing that the intron length varied among different loci. Four different cDNA sequences were identified from one homozygous diploid Japanese flounder individual, implying the existence of at least four loci. Comparison of the cDNA sequences to the DNA sequence confirmed that six exons existed in this gene of Japanese flounder, which was a common feature shared by Pleuronectiformes fishes. Our results proved the multi-locus feature of MHC class II B. The sequences we obtained would provide detailed and systematic data for further research.