细粒棘球绦虫核糖体蛋白S9基因克隆鉴定及其在不同发育阶段表达分析  

Cloning and analysis of the ribosomal protein S9 gene from Echinococcus granulosus and analysis of its expression in different stages of Echinococcus granulosus development

作　　者：肖云峰[1,2,3] 刘辉[3] 吕国栋[3] 赵军[3] 高惠静[3] 李娟[3] 王建华[1,2,3] 

机构地区：[1]新疆医科大学 [2]新疆医科大学基础医学院,新疆乌鲁木齐830054 [3]新疆医科大学第一附属医院

出　　处：《中国病原生物学杂志》2014年第2期150-154,共5页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.81000746);新疆医科大学第一附属医院科研专项基金资助项目(No.2010YX01)

摘　　要：目的克隆鉴定细粒棘球绦虫原头蚴核糖体蛋白S9基因，分析其在细粒棘球绦虫不同发育阶段的表达情况。方法从已构建的cDNA文库挑选基因，克隆EgRPS9基因并对编码产物的理化性质、功能位点、蛋白质的结构和功能域进行预测和分析，通过Real Time-PCR检测RPS9基因在细粒棘球绦虫不同发育阶段的表达情况。结果通过各种生物数据库对EgRPS9基因进行预测并测序，显示该基因具有完整开放阅读框，大小为564bp，编码188个氨基酸，预测分子质量单位为22．0ku，等电点为10．32。SMART保守功能区域分析发现该基因第107～149个氨基酸为40s亚基即结构域s4超家族。通过多序列比对发现该基因具有较高的保守性，进化树结果表明该基因与曼式血吸虫、日本血吸虫及华氏睾吸虫亲缘关系较近。RealtimePCR检测原头蚴和囊泡RPS9基因相对表达量分别为1．16和1．02，差异无统计学意义（P〉0．05）。结论成功克隆出细粒棘球绦虫40S亚基RPS9新基因，该基因在细粒棘球绦虫不同发育阶段均有表达。Objective To clone and identify the ribosomal protein S9 gene from Echinococcus granulosus and analyze its expression in different stages of E. granulosus development. Methods The EgRPS9 gene was cloned from a cDNA li brary of E. granulosus, and its sequence was analyzed using bioinformatics. The physicochemical properties, functional sites, structure, and function domains of proteins were predicted and analyzed. Relative quantitative PCR with 2 standard curves was used to confirm the quantity of EgRPS9 gene expression in different stages of E. granulosus development. Results Bioinformatic analysis revealed that the EgRPS9 gene bad a complete open reading frame of 564 bp encoding 188 amino acids. The EgRPS9 protein had a molecular weight of 22.0 ku and an isoelectric point of 10.32. SMART database analysis revealed a ribosome 40s subunit S4 super family domain located at amino acids 107--149. Multi-sequence align ment revealed that tiffs gene is highly conserved. A phylogenetie tree revealed that this gene is closely related to Schistoso ma mansoni , Schistosoma japonicum , and Clonorchis sinensis. Results of real-time PCR revealed that the quantity of gene expression was 1.16 and 1.02 in different stages of E. granulosus development. Differences in the quantity of gene ex pression were not statistically significant （P〉0. 05）. Conclusion A new gene of the ribosome 40s subunit, RPSg, from E. granulosus was successfully cloned. This gene is expressed in E. granulosus metacestodes.

关 键 词：细粒棘球绦虫 核炙糖体蛋白S9基因 实时荧光定量PCR 基因克隆 序列分析 

分 类 号：R383.33[医药卫生—医学寄生虫学]