青海血蜱cDNA表达文库的免疫学筛选和阳性克隆的鉴定  

Immunoscreening of cDNA Expression Library of Haemaphysalis qinghaiensis and Characterization of Positive Clones

作　　者：高金亮[1] 罗建勋[1] 樊瑞泉[1] 魏永红 李文卉[1] 任巧云[1] 关贵全[1] 殷宏[1] 

机构地区：[1]中国农业科学院兰州兽医研究所,甘肃兰州730046 [2]甘肃省临潭县畜牧中心,甘肃临潭747500

出　　处：《中国兽医学报》2006年第6期631-633,共3页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(30270992);国家"863"计划资助项目(2003AA241110)

摘　　要：为获得抗青海血蜱免疫原基因,用兔抗青海血蜱差异蛋白阳性血清和兔抗青海血蜱唾液蛋白阳性血清对青海血蜱cDNA表达文库进行了免疫学筛选,经过初筛和复筛共获得58个阳性信号。用所得阳性噬菌体转染宿主菌BM25.8使之自动亚克隆为重组质粒,用此亚克隆质粒转化宿主菌JM109并从中提取重组质粒进行PCR、酶切和测序分析。序列分析表明:共获得新cDNA序列21个。将前5个cDNA登录GenBank/ncbi,获取登录号。所获阳性克隆为青海血蜱保护性抗原的筛选奠定了基础。：To obtain the immunogen against Haemaphysalis qinghaiensis,a cDNA expression library of H. qinghaiensis was screened with rabbit anti-H, qinghaiensis differential protein serum and rabbit anti-H, qinghaiensis saliva protein serum. Totally 58 positive signals were obtained. Pure phage stocks were harvested by plaque purification and converted to plasmid subclones by plating phage on host strain BM25.8. Recombinant plasmids that were subcloned in BM25.8 were isolated and then transformed into host strain JM109. Twenty one positive new clones were obtained after analysis of plasmids abstracted from JM109 by PCR,restriction digestion and sequencing. Five of the new clones were submitted to the GenBank/DDBJ/EMEL nucleotide sequence database.

关 键 词：青海血蜱 CDNA表达文库 免疫学筛选 

分 类 号：S852.746[农业科学—基础兽医学]