刺毛虫毒刺cDNA文库的构建及初步分析  

Construction and preliminary analysis of a cDNA library from the spines of the slug moth(Lepidoptera:Eucleidae)

作　　者：李滨[1] 欧阳碧函[2] 徐宏博[2] 罗玉娇[1] 蒋立平[1] 舒衡平[1] 

机构地区：[1]中南大学基础医学院病原生物学系,湖南长沙410078 [2]中南大学湘雅医学院临床医学八年制

出　　处：《中国病原生物学杂志》2011年第11期828-830,836,F0004,共5页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.30901874);中南大学中央高校基本科研业务费专项(No.2010QZZD028),中南大学研究生学位论文创新基金资助(No.2010ssxt260),中南大学贵重仪器设备开放共享基金项目(No.2010-018);大学生创新性实验计划湖南省级项目(No.YB10074)

摘　　要：目的构建刺毛虫毒刺的cDNA文库,克隆其毒素编码的基因。方法提取刺毛虫毒刺中的总RNA;使用clontech公司的cDNA文库构建试剂盒进行反转录,长距离PCR方法合成第二条链,酶切纯化后与载体连接,电转化入感受态细胞XL1-Blue,通过抗性筛选后进行菌落PCR鉴定,挑选插入子在300bp以上的单克隆送至测序公司测序。结果成功构建了刺毛虫毒刺cDNA文库,初步鉴定其插入子长度为250～1 000bp。结论刺毛虫毒刺cDNA文库构建成功,并获得大量的编码刺毛虫毒素的全长基因,为研究其致病机制奠定了基础。Objective To construct a cDNA library of a gene coding for toxin in the spines of the slug moth（Lepidoptera： Eucleidae）.Methods Total mRNA was extracted from the slug moth＇s spines using a SMARTTM cDNA Library Construction Kit from Clontech Laboratories.The second chain was replicated,linked to the vector,and then transformed into XL1-Blue competent cells.Mono-clones with inserts larger than 300 bp were sequenced after screened with PCR.Results The cDNA library was successfully constructed.Target fragments 250-1 000 bp in size were preliminarily identified.Conclusion A cDNA library of a gene coding for the toxin in the spines of a slug moth was successfully constructed,and the entire sequence of the gene coding for this spine toxin was obtained.This should provide a basis for future research on the mechanism of the toxin＇s pathogenicity.

关 键 词：刺毛虫 毒素 CDNA文库 构建 

分 类 号：R384[医药卫生—医学寄生虫学]