重组犬瘟热病毒F-H融合基因乳酸菌的构建及表达

为了构建重组犬瘟热病毒(CDV)F-H融合基因工程乳酸菌,采用重叠延伸PCR(SOE-PCR)技术扩增CDV F-H融合基因。将F-H融合基因亚克隆至穿梭载体p SIP409多克隆位点上,构建p SIP-F-H表达重组子,并电转化至植物乳杆菌NC8感受态细胞中。利用SDS-PAGE和Western blotting检测F-H融合蛋白的表达情况。结果表明,成功地扩增了CDV F-H融合基因,构建了p SIP-F-H重组表达质粒,并电转化至乳酸杆菌感受态细胞中。SDS-PAGE和Western blotting检测表明,重组乳酸杆菌表达了分子量约为62.96 ku的F-H融合蛋白,且该蛋白能与CDV阳性抗体反应,具有反应原性。

Construction and expression of recombinant Lactobacillus containing F-H fusion gene of canine distemper virus from minkSU Feng-yan1,LI Zhe1,LI Yan-zhi3,WANG Cun-feng2,ZENG Fan-li1

In order to construct Lactobacillus containing a F-H fusion gene of canine distemper virus(CDV),the F-H fusion gene was amplified by gene splicing by overlap extension PCR(SOE-PCR).The F-H fusion gene was sub-cloned into shuttle carrier pSIP409 to construct a recombinant plasmid(named as pSIP-F-H).Then the recombinant plasmid pSIP-F-H was elec-transformed into Lactobacillus plantarum competence cells. SDS-PAGE and Western-blot were used to detect the expression of F-H fusion protein. In results, the F-H fusion gene of CDV was amplified successfully.The recombinant plasmid pSIP-F-H was constructed and elc-transformed into Lactobacillus plantarum competence cells successfully. Results of SDS-PAGE and Western-blot showed that recombinant Lactobacillus expressed a fusion protein(nearly 62.96ku in size) and the protein can react with CDV positive antibody.