麦二叉蚜体内病毒结合蛋白基因的克隆和原核表达  

Cloning and Prokaryotic Expression of Virus Binding Protein Gene from Schizaphis graminum

作　　者：吴云锋[1] 林林[1] 崔晓峰[1] 

机构地区：[1]西北农林科技大学植物保护学院,陕西杨凌712100

出　　处：《病毒学报》2002年第3期275-279,共5页Chinese Journal of Virology

基　　金：国家自然科学基金 ( 3 9970 483 ) ;国家八六三青年基金( 97-13 7)

摘　　要：从麦二叉蚜体内克隆了一个DNA片段 ,经序列测定表明该片段全长为 16 4 7bp ,编码 5 4 8个氨基酸。与禾谷缢管蚜的病毒结合蛋白基因核苷酸同源性为 92 % ,氨基酸的同源性为 96 % ,从而认为这是病毒结合蛋白基因(GenBank登录号为AF4 34719)。构建了该基因的原核表达载体 pBVSG和 pETSG ,用pBVSG表达出 6 3kD的非融合目的蛋白 ,用pETSG表达出 6 9kD的融合蛋白 ,二者均有较高的表达量。以纯化的融合蛋白免疫家兔 ,制备了此病毒结合蛋白的抗血清 ,用琼脂双扩散法测定效价为 1∶5 12。A sequence of DNA cloned from Schizaphis graminum by PCR technique contains 1647bp,which can encode 548 amino acids Homology of nucleotide and amino acid sequences in comparison with virus binding protein(VBP) gene of Rhopalosiphum padi were 92% and 96% respectively,so this gene was identified as virus binding protein gene of Schizaphis graminum(GeneBank accession number is AF434719) The cloned VBP gene of Schizaphis graminum were ligated to pVB221,pET 30a and pTrcHisA vectors for expressing The results revealed that the 63kD protein could be expressed using pBV221 When pET-30a was used,the expressed fusion protein had a molecular weight of 69kD,and both of them had a high expression level The fusion protein was purified and was used to immunize rabbit to raise the antibody against VBP Titer of antiserum was 1:512 by double agar diffusion

关 键 词：麦二叉蚜 病毒结合蛋白 基因 克隆 原核表达 小麦黄矮病 

分 类 号：S435.121.4[农业科学—农业昆虫与害虫防治]