鸡MHCⅠ类分子克隆及其二聚体融合基因的构建与表达  

Cloning and Fusion Expression of Chicken MHCⅠ Molecule

作　　者：戴银[1] 程宝艳[1] 胡晓苗[1] 王骏俊[2] 陈芳芳[2] 余为一[2] 

机构地区：[1]安徽省农业科学院畜牧兽医研究所,安徽合肥230031 [2]安徽农业大学动物科技学院,安徽合肥230036

出　　处：《中国畜牧兽医》2011年第3期61-64,共4页China Animal Husbandry & Veterinary Medicine

基　　金：安徽省科技攻关项目(04013042)

摘　　要：为进一步研究MHCⅠ类分子作用机制,以及为制备MHCⅠ基因工程疫苗奠定基础,克隆鸡MHCⅠ基因,并构建MHCⅠβ2m-α融合基因进行原核表达。运用RT-PCR方法,克隆鸡MHCⅠα和β2m链基因,并通过编码连接肽(Gly4Ser)4的基因序列将两者头尾相连,构建了pET-32a-MHCⅠβ2m-α重组质粒,经PCR扩增、双酶切和测序鉴定后,重组质粒在E.coliBL21细胞进行IPTG诱导表达融合蛋白,采用SDS-PAGE和Western blotting方法分别检测表达产物。结果表明,成功克隆了MHCⅠα和β2m链基因,长度分别为1014和300 bp;构建的融合基因MHCⅠβ2m-α长度为1194 bp,经原核表达,融合蛋白分子质量约为62.0 ku,能与相应的抗体结合,具有一定的免疫学活性。To explore MHC mechanism presenting antigen and function as vaccine carrier,we cloned MHCⅠgene from chicken by RT-PCR,and constructed a recombined plasmid containing MHCⅠβ2m-α fusion gene and expressed in a prokaryotic system.Chicken MHCⅠα and β2m chains were cloned from chicken by RT-PCR,the recombinant gene MHCⅠβ2m-α was linked with a DNA sequence coding for short peptide（Gly4Ser）4,and confirmed by PCR amplification,double restriction digestion and DNA sequencing.The recombinant plasmid was expressed in E.coli BL21 after induction by IPTG.SDS-PAGE and Western blotting were used to detect expression products.The results revealed that the lengths of MHCⅠα and β2m chains were 1014 and 300 bp,respectively,and the recombinant fusion gene was 1194 bp.The fusion protein was about 62.0 ku and had the reactinogenicity with specific antibody.

关 键 词：鸡MHCⅠα链 鸡MHCⅠβ2m链 融合蛋白 

分 类 号：Q78[生物学—分子生物学]