期刊文献+
任意字段
题名或关键词
题名
关键词
文摘
作者
第一作者
机构
刊名
分类号
参考文献
作者简介
基金资助
栏目信息

pIRES2-HPV18E2-EGFP质粒重组及其在HeLa细胞中的表达 被引量：1

Construction of recombinant plasmid PIRES2-HPV18-EGFP and its expression in HeLa cells
下载PDF
导出
摘要 目的构建人乳头瘤病毒18型E2基因片段(HPV18E2)重组表达质粒并予以表达,为进一步研制HPV18相关疾病提供材料。方法以重组质粒(pBR322-HPV18)为模板,PCR方法扩增HPV18E2DNA片段,将HPV18E2DNA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。RT-PCR鉴定转染细胞中E2基因。结果PCR扩增DNA片段约为1 kb,与预期结果相同。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。转染细胞RT-PCR出现特异性条带。结论成功构建表达HPV18 E2蛋白的靶细胞模型。 Objective To construct a recombinant plasmid for expressing the HPV18 E2 gene and develop some materials for HPV18 related disease. Methods The E2 gene of HPV18 was amplified by PCR from PBR322-HPV18 and cloned into PIRES2-EGFP. The sequence of cloned HPV18E2 was confirmed by restriction analysis and DNA sequencing, then transfected into HeLa cells. E2 gene was detected by RT-PCR. Results HPV18 E2 gene fragment was amplified by PCR and ligated to PIRES2-EGFP, then transfeeted into HeLa cells successfully. The expression of green fluorescence cells was observed by fluorescence microscopy. Specific band could be seen in transfeeted cells by RT-PCR. Conclusion Recombinant plasmid PIRES2-EGFP is effectively expressed after being transfected into HeLa cells in vitro.
作者 汪云 李力 李元朝 吕凤林
机构地区 第三军医大学大坪医院野战外科研究所妇产科 第三军医大学大坪医院野战外科研究所第一研究室创伤烧伤与复合伤研究室
出处 《第三军医大学学报》 CAS CSCD 北大核心 2006年第12期1280-1282,共3页 Journal of Third Military Medical University
关键词 人乳头瘤病毒18型 E2基因 基因重组 转染 human papillomavirus type 18 E2 gene plasmid construction gene transfection
分类号 R394-33 [医药卫生—医学遗传学] R394.2 [医药卫生—医学遗传学]
  • 相关文献

参考文献6

  • 1THIERRY F,BENOTMANE M A,DEMERET C,et al.A genomic approach reveals a novel mitotic pathway in papillomavirus carcinogenesis[J].Cancer Res,2004,64(3):895-903.
  • 2HUANG L W,CHAO S L,CHEN P H,et al.Multiple HPV genotypes in cervical carcinomas:improved DNA detection and typing in archival tissues[J].J Clin Virol,2004,29(4):271-276.
  • 3IM S S,WILCZYNSKI S P,BURGER RA,et al.Early stage cervical cancers containing human papillomavirus type 18 DNA have more nodal metastasis and deeper stromal invasion[J].Clin Cancer Res,2003,9(11):4145-4150.
  • 4DEFILIPPIS R A,GOODWIN E C,WU L,et al.Endogenous human papillomavirus E6 and E7 proteins differentially regulate proliferation,senescence,and apoptosis in HeLa cervical carcinoma cells[J].J Virol,2003,77(2):1551-1563.
  • 5DEMERET C,GARCIA-CARRANCA A,THIERRY F.Transcription-independent triggering of the extrinsic pathway of apoptosis by human papillomavirus 18 E2 protein[J].Oncogene,2003,22(2):168-175.
  • 6BRANDSMA J L,SHLYANKEVICH M,ZHANG L,et al.Vaccination of rabbits with an adenovirus vector expressing the papillomavirus E2 protein leads to clearance of papillomas and infection[J].J Virol,2004,78(1):116-123.

同被引文献12

  • 1马英,于成银,冷志,邓仁洪,林明杰.HPV感染与宫颈鳞状上皮内病变及宫颈癌关系探讨[J].重庆医学,2004,33(7):1011-1013. 被引量：11
  • 2Bray F, Sankila R, Ferlay J, et al. Estimates of cancer incidence and mortality in Europe in 1995[J]. EJC, 2002, 38:99.
  • 3Vizcaino AP, Moreno V, Bosch FX, et al. International trends in the incidence of cervical cancer: I .Adenocarci- noma and adenosquamous cell carcinomas[J].lnt J Cancer,1998,75:536.
  • 4Resnick RM, Cornelissen MTE, Wright DK, et al. Detection and typing of human papillomavirus in archival cervical cancer specimens by DNA amplification with consensus premers[J]. J Natl Cancer Inst, 1990,82 : 1477.
  • 5Clifford GM, Smith JS, Aguado T, et al. Comparison of HPV type distribution in highgrade cervical lesions and cervical cancer : a meta-abalysis[J]. Br J Cancer, 2003,89 101.
  • 6Albers A, Abe K, Hunt J. Antitumor activity of human papillomavirus type 16,18 ET-specifie T cells against virally infected squamous cell carcinoma of the head and neck[J].CancerRes,2005,65:11146.
  • 7Thierry F, Benotmane MA, Demeret C, et al. A genomic approach reveals a novel mitotic pathway in papillomavirus carcinogenesis[J]. Cancer Res, 2004,64 : 895.
  • 8Im SS,Wilczynski SP,Burger RA,et al. Early stage cervical cancers containing human papillomavirus type 18 DNA have more nodal metastasis and deeper stromal invasion [J]. Clin Cancer Res,2003,9:4145.
  • 9Francis DA, Schmid SI, Howley PM. Repression of the integrated papillomavirus F.6/E7 promoter is required for growth suppression of cervical cancer cells[J]. J Virol, 2000,74:2679.
  • 10DeFilippis RA, Goodwin EC, Wu L, et al. Endogenous human papiltomavirus E6 and E7 proteins differentially regulate proliferation, senescence, and apoptosis in HeLa cervical carcinoma celts[J]. J Virol, 2003,77 : 1551.

引证文献1

二级引证文献1

第三军医大学学报
2006年 第12期

相关作者

内容加载中请稍等...

相关机构

内容加载中请稍等...

相关主题

内容加载中请稍等...

浏览历史

内容加载中请稍等...
;
使用帮助 返回顶部