RACE技术扩增及克隆中华菊头蝙蝠ACE2基因  

Amplification and cloning of angtiotension-coverting enzyme 2 gene from Rhinolophus sinicus by RACE Technique

作　　者：高俊[1] 徐霞[1] 肖洪广[1] 谢天赐[1] 林勇平[1] 

机构地区：[1]广州医学院第一附属医院检验科,广东广州510120

出　　处：《中国热带医学》2009年第4期602-604,共3页China Tropical Medicine

基　　金：广东省自然科学基金面上项目(06022094);广州市教委青年科研基金(1047)

摘　　要：目的建立中华菊头蝠体内扩增血管紧张素转换酶2(ACE2)基因真核表达载体,研究SARS冠状病毒感染的跨种属传播机制。方法利用基因的种属保守性分别设计用于扩增5’未端和3’端cDNA的引物,从中华菊头蝠小肠组织中提取总RNA,在通过cDNA末端快速扩增(RACE)技术获得cDNA 5’端和3’端序列基础上,进一步PCR扩增ACE2开放阅读框并克隆到pcDNA3.1。结果成功获得了中华菊头蝠的ACE2基因全长序列,并构建了其真核表达载体pcDNA3.1/R-ACE2。结论利用RACE技术扩增并构建了中华菊头蝙蝠的ACE2的真核表达载体,为研究SARS冠状病毒感染与免疫及跨种属传播机制奠定了基础。Objective To amplify and clone the coding region for the angtiotension-coverting enzyme2（ACE2） in the bat of Rhinolophus sinicus using RACE technology. Methods Primers for amplified 3＇-cDNA and 5＇-cDNA were designed according to the multi -alignment analysis of ACE2 Coding sequencing from different mamalian species.Total RNA was extracted from intestine tissue and ACE2 gene of Rhinonophus sinicus was amplified by RACE.PCR product was cloned into eukaryotic expression plasmid vector pcDNA3.1 （＋） and comfirmed with sequencing. Results Bat ACE2 encode gene was successfully amplified and its eukaryotic expression plasmid was constructed. Conclusion Amplification and cloning of bat- ACE2 could provide the basis for further investigation on the mechanism of interspecies transmission of SARS-CoV.

关 键 词：CDNA末端快速扩增 血管紧张素转换酶2 蝙蝠 

分 类 号：R373.1[医药卫生—病原生物学]