抗白念珠菌人源性单链抗体库的构建及筛选  

Construction and Screening of Human Single-chain Antibody Library for Candida albicans

作　　者：路艳[1] 韩跃武[1] 韩亚萍[1] 刘玲玲[1] 王春霞[1] 李真真[1] 

机构地区：[1]兰州大学基础医学院生物化学与分子生物学研究所,兰州730000

出　　处：《中国生物制品学杂志》2009年第11期1063-1067,共5页Chinese Journal of Biologicals

基　　金：国家"863"计划重大项目子课题(2006AA10A208-1-4);甘肃省科技攻关项目(2GS064-A43-020-21)

摘　　要：目的构建多样性良好的抗白念珠菌人源性单链抗体库,筛选抗白念珠菌特异性的噬菌体抗体。方法从20份人外周血淋巴细胞(包括正常成年人5份、新生儿5份和白念珠菌感染恢复期患者10份)中提取总RNA,反转录为cDNA,PCR扩增人抗体重链(VH)和轻链(VL)可变区基因,以重叠延伸PCR法将VH和VL拼接成scFv基因,克隆入噬菌粒载体pCANTAB-5E中,电转化大肠杆菌XL1-Blue,构建人源抗白念珠菌天然噬菌体抗体库,并从中筛选阳性克隆抗体。结果构建的人源性抗白念珠菌天然噬菌体抗体库库容为2.8×109,多样性良好。共筛选出18个阳性克隆。结论已成功构建了1个多样性良好的抗白念珠菌人源性噬菌体抗体库,为筛选有效治疗白念珠菌和耐药性白念珠菌感染的药物提供了条件。Objective To construct a human single-chain antibody （scFv） library for Candida albicans with diversity. Methods Total RNAs were extracted from peripheral blood lymphocytes of 20 persons, including 5 healthy adults, 5 newborns and 10 patients with Candida albicans infection in recovery phase, and transcribed to cDNA for amplification of V, and V1 genes by PCR. The amplified V. and Va genes were spliced to form scFv gene which was cloned into phagemid pCANTAB-SE, and the construcled recombinant phagemid was transformed to E. coli XL1-Blue to construct human natural single-chain antibody library from which positive clones were screened. Results A human natural single-chain antibody library with capacity of 2. 8 × 10^9 and diversity was constructed, from which 18 positive clones were screened. Conclusion A human natural single-chain antibody library for Candida （dbicans with diversity was successfully constructed, which prnvided a condition for screening of drugs against infection with Ccmdida albicans or drug-resistant Candida albicans.

关 键 词：白念珠菌 天然抗体库 单链抗体 

分 类 号：Q782[生物学—分子生物学] Q81