白纹伊蚊AL-100 30ku蛋白基因的克隆及其原核表达载体的构建  

Cloning and prokaryotic expression vector construction of AL-100 30ku gene from Aedes albopictus

作　　者：赵郭存[1] 刘志刚[1] 邬玉兰[1] 孔小丽[1] 

机构地区：[1]深圳大学医学院过敏反应与免疫学研究所,广东深圳518060

出　　处：《中国病原生物学杂志》2009年第6期429-431,443,共4页Journal of Pathogen Biology

基　　金：国家863项目(No.2006AA10Z236);广东省科技重点专项资助项目(No.2003A3080502)

摘　　要：目的克隆白纹伊蚊主要过敏原AL-100 30ku蛋白基因,并构建其原核表达载体。方法RT-PCR克隆白纹伊蚊主要过敏原AL-100 30ku的全长基因,设计简并引物,扩增白蚊伊蚊30ku的完整开放阅读框,与pET-28a载体连接,构建原核表达载体。结果成功克隆白纹伊蚊主要过敏原30ku基因并构建其原核表达载体。该基因含有长度为816bp的开放阅读框,编码271个氨基酸。该蛋白质的分子质量单位为28.33ku,等电点为4.04,编码序列与数据库中已知的白纹伊蚊30ku基因的同源性为99%。结论成功克隆了白蚊伊蚊主要过敏原30ku基因并构建了原核表达载体,为白纹伊蚊主要过敏原30ku蛋白的重组表达和免疫活性鉴定等奠定了基础。Objective Cloning and prokaryotic expression vector construction of AL-100 30 ku gene from Aedes albopictus. Methods The RT-PCR was applied to clone the full-length genes of major allergen AL-100 30 ku from Ae. albopictus and then the sequences were analyzed. The specific primers were designed. The ORF of AL-100 30 ku of Ae. albopictus was subcloned into the expression vector pET 28a. Results AL-100 30 ku gene from Ae. albopictus was cloned and constructed to prokaryotic expression vector. The cloned cDNA ORF sequence contained 816 bp and encoded 271 amino acids. Sequence analysis showed that the estimated molecular mass of 28 330, pI4.04, the cloned sequence homology was high to 99% with AL-100 30 ku from Ae. albopictus. Conclusion AL-100 30 ku major allergen gene from Ae. al-bopictus was cloned and constructed to the prokaryotic expression vector successfully. It establishes the foundation for the recombinant allergen of 30 ku protein expression and the activity identification.

关 键 词：白纹伊蚊 过敏原 AL-100 30 KU 克隆 原核表达载体 

分 类 号：R384.1[医药卫生—医学寄生虫学]