期刊论文

A Construction of Eucaryotic Expression Vector for Outer Membrane Protein Tp92 from Treponema Pallidum and its Expression in Hela Cells

中文

实用预防医学

1006-3110

2005

12

1

9-13

10.3969/j.issn.1006-3110.2005.01.003

湖南省教育厅重点资助项目 （0 0 2A0 0 8）； 湖南省卫生厅科研基金 （B2 0 0 3 - 0 85）；

本校为第一机构

目的以梅毒螺旋体(Treponema pallidum,Tp)外膜蛋白基因Tp92为目的基因,构建Tp真核表达重组体pcDNA3.1(+)-Tp92,并鉴定其在Hela细胞能否正确表达,为进一步开展TpDNA疫苗动物实验打下基础.方法应用PCR技术从Tp Nichols株基因组模板中扩增Tp92基因,定向克隆构建真核表达重组体pcDNA3.1(+)-Tp92,脂质体介导将pcDNA3.1(+)-Tp92转染入Hela细胞,以免疫酶标法和一步法RT-PCR检测pcDNA3.1(+)-Tp92在Hela细胞中的表达情况.结果双酶切及测序鉴定证明成功构建Tp真核表达重组体pcDNA3.1(+)-Tp92,DNA测序显示重组质粒含有2 103 bp的目的基因片段,读码框架正确,无碱基错配及移码突变.测序后结果经与GenBank登录...

Objective To construct the recombinant plasmid of Eucaryotic expression containing Tp92 Gene from Treponema pallidum,and transfect it into Hela cells to express the outer membrane protein. Methods Tp92 Gene was amplified from the genomic DNA of T. pallidum by polymerase chain reaction (PCR),and the gene was inserted into cloning vector pUCm-T.The inserted Tp92 gene was subcloned into appropriate site of pcDNA3.1(+) vector.After identification by sequencing and restrictive enzyme digestion,the recombinant plasmid was transfected into Hela cells ...