黄颡鱼MHC classⅡ基因全长的克隆及饲料维生素D3对其组织表达的影响

FULL-LENGTH cDNA CLONING AND THE EFFECT OF DIETARY VITAMIN D_3 ON TISSUE EXPRESSION OF MAJOR HISTOCOMPATIBILITY COMPLEX-Ⅱ IN YELLOW CATFISH(PELTEOBAGRUS FULVIDRACO)

为进一步丰富鱼类MHC class Ⅱ基因的研究,同时也为进一步探讨低磷饲料中添加维生素D3对鱼类免疫功能可能的影响,实验利用RACE(Rapid-amplification of c DNA ends)即c DNA末端快速扩增技术,成功克隆出黄颡鱼(Pelteobagrus fulvidraco)主要组织相容性复合体(Major histocompatibility complex,MHC)class Ⅱ抗原基因,全长1074 bp,其中ORF(Open reading frame)708 bp,编码236个氨基酸,5′UTR(5′端非翻译区)78 bp,3′UTR(3′端非翻译区)259 bp。进行氨基酸序列比对分析得到:黄颡鱼MHC class Ⅱ基因ORF氨基酸序列与长吻逘(Leiocassis longirostris)的氨基酸序列相似度最高为69.5%,与锦鲤(Cyprinus carpio)的氨基酸序列相似度最低为50.4%。利用q PCR对黄颡鱼MHC class Ⅱ基因进行组织表达分析,结果表明MHC class Ⅱ在小肠、肝脏、鳃中表达较高;在肌肉、鳍条中表达较低;而在肾、脾脏、脑、头肾中表达量极低(几乎检测不到)。在低磷饲料中添加维生素D3显著诱导了该基因的上调表达。研究结果展示了黄颡鱼MHC class Ⅱ抗原基因的分子结构、组织表达以及维生素D3的作用,在降低磷排放的同时,为今后黄颡鱼免疫抗病及分子选育等方向的深入研究及免疫型饲料的使用奠定了基础。

To study the relationship between vitamin D3(VD3)and the major histocompatibility complex(MHC)classII gene,the MHC class II antigen gene of yellow catfish was cloned by rapid-amplification of cDNA ends.The fulllengthcDNA of MHC class II is1074bp long and comprises a78bp5′untranslated region(UTR_,259bp3′UTR,anda708bp open reading frame that encodes236amino acids.The deduced amino acid sequence showed the highest identity(69.5%)and the lowest identity(50.4%)with Leiocassis longirostris and Cyprinus carpio,respectively.Expressionof the MHC class II antigen gene was relatively high in the small intestine,liver,and gill,but low in muscles and tail.Itwas nearly undetectable in the other tissues.Vitamin D3(0,5000IU/kg)was added to low phosphorus diets in thegrowth trial.The addition of dietary VD3significantly stimulated MHC class II gene expression in the small intestineand kidney.This study describes the molecular structure,tissue expression of the MHC class II antigen gene,and possibleeffects of dietary VD3on tissue expression of the MHC class II gene in yellow catfish,providing useful informationfor further studies of the function of this gene in yellow catfish.