犬细小病毒VP2基因在枯草芽孢杆菌中的表达及鉴定  

Expression and Characterization of Canine Parvovirus VP2 Gene in Bacillus subtilis

作　　者：李爽[1] 黄文斌[1] 黄爱玲[1] 吕暾[1] 

机构地区：[1]福州大学生物科学与工程学院,福州350108

出　　处：《生物技术通报》2011年第12期171-174,共4页Biotechnology Bulletin

摘　　要：从犬细小病毒/犬瘟热二联苗中提取CPV基因组,根据GenBank发表的CPV-VP2基因序列设计一对引物,对VP2基因进行PCR扩增,并克隆至TA Cloning Kit Dual Promoter(pCRⅡ),获得克隆载体pCR-VP2。将重组质粒亚克隆到枯草芽孢杆菌表达载体pHT43上,获得重组表达载体pHT43-VP2。经双酶切鉴定及序列比对分析后,将pHT43-VP2载体转化入枯草芽孢杆菌WB600中进行诱导表达,产物使用PAGE胶蛋白回收法纯化。结果表明,在69 kD处存在目的蛋白,ELISA检测发现,纯化后的目的蛋白与阳性血清存在特异性反应。The genome of CPV was extracted from CPV/CDV vaccine.A pair of primers was designed according to the sequence of CPV-VP2 gene from GenBank.Polymerase chain reaction（PCR） was used to amplify the VP2 gene.Then the fragment was cloned into TA Cloning Kit Dual Promoter（pCRⅡ） to obtain a cloning vector pCRⅡ-VP2.Then the recombinant plasmid was subcloned into the expression vector pHT43 named pHT43-VP2.The pHT43-VP2 was verified by restriction enzymes and nucleotide sequence analysis.The expression vector was transformed into Bacillus subtilis and the expressed product was recovered from PAGE gel.The results showed that VP2 protein expressed has 69 kD of molecular weight.ELISA revealed that the protein expressed in B.subtilis can be recognized by the positive serum of CPV.

关 键 词：犬细小病毒 VP2蛋白 枯草芽孢杆菌 

分 类 号：S852.65[农业科学—基础兽医学]