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HPV18E2蛋白CTL表位多肽的构建与表达
引用本文:汪云,肖岚,吕凤林. HPV18E2蛋白CTL表位多肽的构建与表达[J]. 局解手术学杂志, 2007, 16(6): 376-378
作者姓名:汪云  肖岚  吕凤林
作者单位:[1]第三军医大学基础医学部组织学与胚胎学教研室,重庆400038 [2]第三军医大学附属大坪医院野战外科研究所第一研究室,重庆400042
摘    要:
目的分析HPV18E2蛋白的CTL表位,合成并纯化这些表位多肽,构建表达HPV18E2蛋白的靶细胞,检验特异性CTL对靶细胞的杀伤效果,为进一步研制人乳头瘤病毒18型(HPV18)基因工程疫苗打下基础。方法以重组质粒(pBR322-HPV18)为模板,利用PCR方法扩增HPV18E2 DNA片段,将HPV18E2 DNA与加强绿色荧光质粒(pIRES2-EGFP)重组构建重组质粒(pIRES2-HPV18E2-EGFP)。用酶切电泳及测序检查质粒重组后序列正确性。重组质粒转染Hela细胞。51^Cr释放实验评估CTL杀伤能力。结果克隆重组质粒pIRES2-HPV18E2-EGFP酶切后显示的酶切图谱与预期相同,而且测序验证插入片段全序列无改变。转染并用G418筛选后,在荧光显微镜下可见绿色荧光细胞的表达。三条候选多肽特异性杀伤能力分别为60.00%、71.29%和80.00%。结论三条备选肽段都具有明显的杀伤效应,均为HPV18E2蛋白的有效CTL表位。

关 键 词:人乳头瘤病毒18型  E2基因  多肽
文章编号:1672-5042(2007)06-0376-03
收稿时间:2007-08-30
修稿时间:2007-10-15

Construction and expression of recombinant plasmid pIRES2-HPV18-EGFP
WANG Yun,XIAO Lan,L Feng-lin. Construction and expression of recombinant plasmid pIRES2-HPV18-EGFP[J]. Journal of Regional Anatomy and Operative Surgery, 2007, 16(6): 376-378
Authors:WANG Yun  XIAO Lan  L Feng-lin
Affiliation:WANG Yun,XIAO Lan,L(U) Feng-lin
Abstract:
Objective To analyse the function of epitope-specifical HPV18E2 CTL. Methods The E2 gene of HPV18 was amplified by PCR from pBR322-HPV18 and cloned into plRES2-EGFP. The sequence of cloned HPV18E2 was confirmed by restriction analysis and DNA sequencing. Then transfected into HeLa cells. The functional analysis is valued by 51^ Cr-release assay. Results HPV18 E2 gene fragment was amplified by PCR and ligated to plRES2-EGFP, then transfected into HeLa cells successfully. The expression of green fluorescence cells was observed by fluorescence microscopy. The specific lysis of the cells from the three peptides was 60.00%, 71.29% and 80.00%. Conclusion The three peptides have the ability to active epitope-specifical CTL. Based on them, a new peptides vaccine maybe constructed.
Keywords:human papillomavirus typel 8    E2 gene    polypeptide
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