传染性法氏囊炎病毒部分VP2基因的克隆与原核表达.pdfVIP

安徽农业大学学报，2008，35(2)：207-210 Journal of Anhui Agricuhural University 传染性法氏囊炎病毒部分VP2基因的克隆与 核表达 程帮照，余为一 ，周丽莎，谢金文，程宝艳 (安徽农业大学动物科技学院，合肥230036) 摘 要：传染性法氏囊炎病毒(IBDV)是侵害雏鸡和青年鸡的重要病原，病毒VP2蛋白是主要保护性抗原。为 研究不同毒株VP2基因的变异及其抗原表位特点，作者克隆、表达和鉴定了VP2基因。首先应用SPF鸡胚接种和 鸡胚成纤维细胞培养法复制病毒，并以自行设计的1对引物，从中扩增出987 bp的VP2片段并进行鉴定。测序结 果表明，国内不同毒株VP2高变区序列同源性达99％以上。将其部分片段插入载体pGEX-4T一1，构建原核表达重 组质粒，转入大肠杆菌后用IPTG诱导表达。经检测，在SDS—PAGE中可见大小为44 400的融合蛋白。 关键词：传染性法氏囊炎病毒；克隆；VP2基因；原核表达 中图分类号：$858．312．65 文献标识码：A 文章编号：1672—352X(2008)02-0207-04 Cloning and expressing of partial VV2 gene of IBDV CHENG Bang—zhao，YU Wei—yi，ZHOU Li—sha，XIE Jin—wen，CHENG Bao—yan (School of Animal Science and Technology，Anhui Agricultural University，Hefei 230036) Abstract：Infectious bursal disease virus(IBDV)was a kind of pathogen that mainly infect young chicken， and its VP2 protein was important protective antigen．This research aims to know the mutation of different IBDV strains and construct prokaryotic expression system．Firstly，IBDVs were proliferated by inoculating SPF chicken embryo or their fibroblast cells respectively．Then a DNA fragment of 987 bp was amplified with a pair of designed primers．After identification，it was inserted into the vector pGEX--4T一1．The result of sequencing indicated that the homology of high variant region of VP2 with strains registered in GenBank is over 99％ ．A part of the fragment was observed from the fragment with another primer to construct prokaryotic expression．Then this fragment was inserted into recombinant plasmid．It was induced in E．coli BI21 by IPTG and identified by SDS—PAGE electrophoresis． The resuh showed that expressed fusion protein was about 44 400．All these suggest that the construction expression system could found for further research of VP2 function． Key words：infectious bursal disea