摘要
制备CVB3结构蛋白和非结构蛋白重组质粒DNA疫苗时 ,采用RT PCR从CVB3感染的HeLa细胞中扩增VP1、VP2、2A和 3D基因 ,重组入真核表达质粒 pcDNA3中 ,构建pcDNA3/VP2、pcDNA3/VP1、pcDNA3/ 2A和pcDNA3/ 3D重组质粒 ,经酶切和测序证实扩增的序列并将各重组质粒体外转染真核细胞COS 7,用RT PCR检测mRNA的转录 ,用Western blot检测表达产物。结果 4种重组质粒酶切出相应大小的目的片段 ,经测序证实为CVB3相应序列 ,Western blot证实能够在体外真核细胞中表达。本文成功构建CVB3结构与非结构蛋白的重组质粒DNA疫苗 ,为进一步研究其免疫效果奠定了基础。
To prepare DNA vaccines of CVB3 structural and nonstructural proteins.VP1、VP2、2A and 3D genes were amplified from CVB3 infected cells with RT PCR and recombinated into eukaryotic expression plasmid,pcDNA3.Four recombinant plasmids were confirmed to contain the amplified target gene segments with restriction digestion and sequencing and they could be transcripted and translated in COS 7 cells in vitro with RT PCR and Western blot.DNA vaccines of CVB3 structural and nonstructural proteins were constructed,which provides the basis on the further study their immune responses in vivo.
出处
《微生物学免疫学进展》
2000年第4期19-22,共4页
Progress In Microbiology and Immunology
