猪囊尾蚴抗原cC1在大肠杆菌中的克隆及高效表达

CLONING AND EXPRESSION OF CYSTICERCUS CELLULOSAE ANTIGEN cC1 IN E.coli

[目的 ]在大肠杆菌中克隆及高效表达猪囊尾蚴抗原cC1。 [方法 ]将cC1cDNA片段以BamHI和PstI克隆到pGEM 3Z载体 ,改换限制性内切酶位点成BamHI和PstI,加上人工接头PstI BamHI XhoI后 ,克隆到pGEX 5T ,构建重组原核表达载体pGE X5T cC1。 [结果 ]培养 3h、IPTG诱导 6h ,cC1表达量最高 ,表达量占细菌总量的 5 7% ,Westernblotting结果表明猪囊尾蚴抗原cC1蛋白与囊尾蚴病猪血清有特异性的结合条带。 [结论 ]猪囊尾蚴抗原cC1在大肠杆菌中获得高效表达。

Objective]To clone and express Cysticercus Cellulosae antigen cC1 in E.coli . [Methods] cC1cDNA fragment was cloned to BamHI and PstI sites of pGEM 3Z vector.After alteration of the restriction sites,the fragment was cloned to EcoRI and XhoI sites of pGEX 5T with a synthetic linker to construct recombinant expression vector pGEX 5T cC1. [Results] The clone produced the largest yield of cC1 protein expression when incubated in 2YT culture medium for 3 h or induced by IPTG for 6 h.Detected by scanning optical densitometry, cC1 constituted 57% of the total bacterial proteins. Western blotting analysis revealed that the GST cC1 fusion protein exhibited a specific reactive band.[Conclusion] High level expression of Cysticercus cellulosae antigen cC1 was obtained in E.coli .