抗脱氧雪腐镰刀菌烯醇噬菌体单链抗体库的构建与鉴定  

Construction and Identification of Phage ScFv Library against Deoxynivalenol

作　　者：周艳红[1] 祭芳[1] 丁衬衬[1] 史建荣[1] 

机构地区：[1]江苏省农业科学院食品质量安全与检测研究所,农业部食品质量安全监控重点开放实验室,江苏省食品质量安全重点实验室,江苏南京210014

出　　处：《江苏农业学报》2010年第5期1093-1097,共5页Jiangsu Journal of Agricultural Sciences

基　　金：国家公益性行业科研专项(3-15);中国博士后科学基金面上资助项目(20080441061);江苏省博士后科研资助计划项目(0901067C);农产品安全检测实验室开放课题(BM2006611)

摘　　要：从抗脱氧雪腐镰刀菌烯醇(DON毒素)杂交瘤细胞株中扩增出VH和VL基因片段,再经重叠延伸PCR拼接扩增得到单链抗体基因(ScFv),克隆到pCANTAB-5E噬菌粒载体上。然后转化感受态大肠杆菌TG1,经辅助噬菌体超感染,构建成抗DON毒素噬菌体单链抗体库,库容约为1.1×105,滴度为1.3×108pfu。随机选取10个克隆进行双酶切鉴定,结果显示均有ScFv基因片段插入。抗DON毒素单链抗体库的构建,为进一步富集筛选并表达抗DON毒素单链抗体奠定了基础。The VH and VL gene fragments were first amplified from cDNA of hybridoma cell line specific to deoxynivalenol,integrated as ScFv with a pair of linker primers,cloned into phagemid vector pCANTAB-5E,and then transformed into Escherichia coli TG1.To produce phage-display ScFv,the helper phage M13KO7 was added to the cultures when bacteria were grown up to the log-phase,in which ScFv antibody was displayed in N-terminus of the bacteriophage coat protein pⅢ.The phage antibody contained 1.1×10^5 independent clones,and the titer of which was about 1.3×10^8 pfu.Enzymatic digestions of 10 random clones showed that the recombinant plasmids contained an intact ScFv gene.These results indicated that the phage antibody library was constructed successfully.

关 键 词：脱氧雪腐镰刀菌烯醇 单链抗体 噬菌体抗体库 

分 类 号：S188[农业科学—农业基础科学]