马属动物cyclinT1基因的扩增与真核表达载体的构建  

Amplification of equine cyclinT1 gene and construction of its eukaryotic expression vector

作　　者：史楠[1] 赵立平[1] 吕晓玲[1] 马建[1] 周建华[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所大动物病研究室兽医生物技术国家重点实验室,黑龙江哈尔滨150001

出　　处：《黑龙江畜牧兽医》2009年第12期1-3,共3页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(30771994)

摘　　要：在慢病毒复制过程中cyclinT1(CycT1)是重要的辅助因子。为了揭示慢病毒的复制机理,试验使用RT-PCR方法扩增马和驴的cyclinT1基因,并进行克隆和序列分析,然后用DNASTAR(MegAlign)软件将测得的序列和国外发表的马属动物CycT1序列(GenBank中登录号为AF137509和AF190905)进行同源性比较。结果表明:马属动物CycT1大小为2184bp,编码727个氨基酸;4种CycT1基因氨基酸同源性在98%以上;将扩增出的马CycT1基因克隆至真核表达载体pcDNA3.1(+)中,经酶切及测序鉴定证实成功构建了pcDNA3.1(+)/eCycT1重组质粒。CyclinT1 is the most important cofactor in lentivirus replication. To reveal the mechanism of the replication, the complete cyelinT1 （ CycT1 ） gene was amplified by RT - PCR from total RNA extracts of equine and donkey ceils, and was cloned and sequenced. The sequencing data indicated that the full lengths of both the equine CyeT1 （ eCycT1 ） and donkey CycT1 （ dCycT1 ） were 2 184 bp, which encoded 727 amino acids. Comparison of the two acquired sequences to the corresponding sequences published on GenBank （ AF137509 for eCycT and AF190905 for dCycT） with DNASTAR demonstrated that the homology in amino acid were over 98%. The ORF of eCycT was subsequently cloned into an eukaryotic expressing vector pcDNA3.1 （ ＋ ）. The resultant pcDNA3.1 （ ＋ ） based eCycT1 expression vector was verified by restriction enzyme analysis and DNA sequencing.

关 键 词：CyclinT1 克隆 序列分析 真核表达载体 

分 类 号：S821[农业科学—畜牧学]