长角血蜱雌蜱唾液腺cDNA表达文库的免疫筛选与初步分析  

Immunoscreening and analysis of cDNA expression library from salivary gland of female Haemaphysalis longicornis

作　　者：刘光远[1] 谢俊仁[2] 田占成[2] 李知新[2] 龚真莉[2] 王路[2] 柴慧萍[2] 才学鹏[2] 

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070 [2]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室甘肃省动物寄生虫病重点实验室,甘肃兰州730046

出　　处：《甘肃农业大学学报》2008年第1期18-24,共7页Journal of Gansu Agricultural University

基　　金：国家科技基础条件平台项目(2005DKA21205-3);国家高新技术研究发展计划“863”项目(2006AA10A207);甘肃省自然科学基金项目(32S041-A25-035)

摘　　要：蜱的免疫预防是目前国内外研究的热点,以抗血清为探针筛选cDNA表达文库是获取功能抗原基因的有效的方法之一,为了获得抗长角血蜱免疫原基因,用兔抗长角血蜱血清和兔抗长角血蜱唾液腺蛋白血清对长角血蜱雌蜱唾液腺cDNA表达文库进行了免疫筛选,经过两轮筛选共获得34个阳性克隆,所得阳性噬菌体转染宿主菌BM25.8使之亚克隆为重组质粒,用此质粒转化宿主菌JM109并进行序列测定和生物信息学分析.结果表明:在长角血蜱雌蜱唾液腺cDNA表达文库中获得26个长角血蜱免疫相关cDNA序列,其编码蛋白与长角血蜱HL35、黏附素hlim3假定蛋白、热休克蛋白、NADH脱氢酶、钙网蛋白以及杂色花蜱和肩突硬蜱的二硫异构酶等具有同源性.所获阳性克隆为长角血蜱保护性抗原基因的筛选奠定了基础.Immunoscreening of cDNA expression library is a useful strategy to obtain the immunogenic genes. Therefore,the cDNA expression library from salivary of female H. longicornis was screened with rabbit anti-H, longicornis serum and rabbit anti-H, longicornis saliva gland serum. After first and second screenings34 positive clones were obtained and transformed into host bacteria JM109 and sequenced. Se-quences analysis indicated 26 unique cDNA sequences that encoded putative proteins showing similarities to HL35 antigen U,hlim3,heat shock protein,NADH dehydrogenase subunit 1 and calreticulin of H. longi-cornis,disulfide isomerase of Amblyomma variegatum and Ixodes scapularis. This will provide the candidates for further screening of protective antigen genes of H. longicornis.

关 键 词：长角血蜱 唾液腺 CDNA表达文库 免疫筛选 

分 类 号：Q789[生物学—分子生物学]