军曹鱼MHC-Ⅰα基因全长cDNA的克隆及其组织表达分析

Full-length cDNA cloning and tissue expression of major histocompatibility complex(MHC)-Ⅰα from cobia(Rachycentron canadum)

采用同源克隆和末端快速扩增（RACE）方法，得到1330bp的军曹鱼（Rachycentroncanadum）MHC-Ⅰα全长cDNA片段。该序列包括76bp的5’末端非编码区（UTR），189bp的3’UTR及1065bp的开放阅读框（ORF），编码354个氨基酸，预测其蛋白质分子量约40．10kDa，等电点5．70。构建MHC-Ⅰα氨基酸序列的系统进化树并进行氨基酸相似性比对，结果表明，军曹鱼和已知鱼类及人类（Homosapiens）MHC-Ⅰα氨基酸的同源性在27．9％～67．1％之间。所推测的蛋白序列具有一些重要特征，包括前导肽、α1、α2、α3区、CP／TM／CYT区和保守的半胱氨酸等。Real—timePCR检测结果显示，MHC-Ⅰα基因在各个正常军曹鱼组织中均表达，但表达量各有不同，其中较强的表达于头肾；中等程度表达于鳃、脾和肠；在心、脑和肌肉中表达较弱。

We cloned the MHC-Ⅰα gene from cobia （Rachycentron canadium） by homology cloning and RACE PCR. The full-length cDNA of MHC-Ⅰα comprises 1 330 bp with a 76 bp 5＇ untranslated region （UTR）, a 189 bp 3＇ UTR and a 1 065 bp open reading frame （ ORF）, encoding a polypeptide of 354 amino acid residues with a predicted molecular weight of 40. 10 kDa and a theoretical iso- electric point of 5.70. According to the phylogenetic tree and amino acid similarity comparison, the homology of MHC-Ⅰα amino acids between cobia and some known fishes and human （Homo sapiens） varies from 27.9% to 67. 1%. The putative protein sequence shows some important features including leader peptide, α1, α2, α3, CP/TM/CYT regions and conserved cysteines, etc.. Real-time PCR result indicates that MHC-Ⅰα expresses in all detected tissues at different expression levels. High expression is detected in head kid- ney; moderate expression is detected in gill, spleen and intestine; while low expression is detected in heart, brain and muscle.