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

HPV16 E6E7与C3d3融合基因真核表达质粒的构建及表达 被引量：2

Construction and expression of HPV16E6E7 and C3d3 fusion gene eukaryotic expression vector
下载PDF
导出
摘要 目的构建可作为DNA疫苗的包含人乳头瘤病毒16型(HPV16)E6E7与C3d3融合基因真核表达质粒,并在体外进行表达和鉴定。方法采用PCR技术扩增HPV16 E6E7片段,将该片段插入到pMDT-18载体中后,亚克隆至真核表达载体pSG.SS.C3d3.YL和pSG.SS.YL,构建E6E7与C3d3融合基因及E6E7表达质粒pSG.SS.E6E7.C3d3.YL和pSG.SS.E6E7.YL。经限制性酶切鉴定和DNA序列测定后,将重组质粒pSG.SS.E6E7.C3d3.YL、pSG.SS.E6E7.YL转染COS-7细胞,用Western blot及免疫细胞化学技术检测其表达。结果经酶切、DNA测序及转染真核细胞后表达产物的鉴定结果显示,重组质粒构建成功。结论构建的质粒可在真核细胞内正确表达,为其作为DNA疫苗诱导免疫效应的动物实验打下了基础。 Objective To construct plasmid pSG. SS. E6E7. C3d3. YL and pSG. SS. E6E7. YL which can respectively express Flk-IECD-C3d3 fusion protein and Flk-IECD protein in eukaryotie cell. Methods PCR was applied to amplifiy E6E7 eDNA with Bgl Ⅱ and BamH Ⅰ restriction endonuclease site at two ends respectively. Then the cDNA fragment was linkaged into pMDT-]8 and was subcloned into eukaryotic expression vector pSG. SS. C3d3. YL and pSG. SS. YL, which leading to pSG. SS. E6E7. C3d3. YL and pSG. SS. E6E7. YL. After identification by endonuclease digestion and DNA sequence analysis, the recombinant plasmids were transfected into COS-7 cells and the expressed product was detected by Western blot and immunocytochemistry. Results The con structions of eukaryotic expression vector pSG. SS. E6E7. C3d3. YL and pSG. SS. E6E7. YL were successful and the two plasmids could express recombinant E6E7-C3d3 fusion protein and Flk-1ECD protein respectively. Conclusion The success in constructions of pSG. SS. E6E7. C3d3. YL and pSG. SS. E6E7. YL do the groundwork for further studies in animals.
作者 郑秀惠 李力 郭建新 郑英如 李军果 林爽 陈竹钦
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妇产科
出处 《重庆医学》 CAS CSCD 北大核心 2010年第1期40-42,共3页 Chongqing medicine
基金 第三军医大学青年基金资助项目(06XG051)
关键词 人乳头瘤病毒16型 C3D3 DNA疫苗 human papillomavirus16 C3d3 DNA vaccine
分类号 R737.33 [医药卫生—肿瘤]
  • 相关文献

参考文献9

  • 1Jung WW,Chun T,Sul D, el al. Strategies against human papillomavirus infection and cervical cancer[J]. J Microbiol,2004,42(4) :255.
  • 2Steller MA. Cervical cancer vaccines: progress and prospects[J]. Soc Gynecol Investig,2002,9(5) :254.
  • 3Rocha Z, avaleta L, Alejandre JE, et al. Parentera and oralimmunization with aplasmid DNA expressing the human papillomavirus16- L1gene induces systemic and rnucosal antibodies and eytotoxic Tlymphoeyteresponses [J]. J Med Virol,2002,66(1) :86.
  • 4Watanabe I, Ross TM,Tamura S, et al. Protection against influenza virus infection by intranasal administration of C3d fused hemagglutinin[J]. Vaccine, 2003, 21 (31): 4532.
  • 5Wang L,Sunyer JO,Bello LJ. Fusion lo C3d enhances the immunogenicity of the E2 glyeoprotein of type 2 bovine viral diarrhea virus[J]. J Virol, 2004,78 (4) : 1616.
  • 6Subramanya D,Grivas PD. HPV and cervical cancer: updates on an established relationship[J]. Postgrad Med, 2008,120(4) :7.
  • 7Sehiffman M,Khan MJ,Solomon D,et al. A study of the impact of adding Human papillomaviruses lypes to cervical cancer screening and triage tests[J]. J Natl cancer Inst, 2005,97(2) : 147.
  • 8Velders MP,Weijzen S, Eiben GL, et al. Defined flanking spacers and enhanced proteolysis is essential for eradication of established tumors by an epitope string DNA vaccine[J]. J Immunol,2001,166 (9) :5366.
  • 9Green TD,Montefiori DC, Ross TM. Enhancement of antibodies to the human immunodeficiency virus type 1 envelope by using the molecular adjuvant C3d[J]. J Virol, 2003,77(3) :2046.

同被引文献18

  • 1Karin H,Jessica M,Frank F,et al. HPV16 E7 oncogene expression in normal human epithelial cells causes molec- ular changes indicative of an epithelial to mesenchymal transition[J]. Virology, 2009,391 ( 1 ) : 57-63.
  • 2Spangle JM,Karl M. The human papillomavirus type 16 E6 oncoprotein activates mTORC1 signaling and increases protein synthesis[J]. J Virol, 2010,84 (18): 9398-9407.
  • 3Yuri A,Koch WM, Xiao WH, et al. Oral HPV infection before and after treatment for HPV16-positive and nega- tive head and neck squamous cell carcinoma [J]. Clin Cancer Res, 2008,14(21) : 7143-7150.
  • 4William L, Schlecht NF, Margaret BG. The role of the hu- man papillomavirus in the pathogenesis of schneiderian in- verted papillomas: an analytic overview of the evidence [J]. Head Neck Pathol,2008,2(2) :49-59.
  • 5Zhou Y, Pan Y, Zhang S, Increased phosphorylation of p70 S6 kinase is associated with HPV16 infection in cer vical cancer and esophageal cancer[J]. Br J Cancer,2007, 97(2) :218 222.
  • 6Dreier K, Scheiden R, Lener B, et al. Subcellular localiza- tion of the human papillomavirus 16 E7 oncoprotein in CaSki cells and its detection in cervical adenocareinoma and adenocarcinoma in situ[J]. Virology, 2011,409 ( 1 ) : 54-68.
  • 7Maelle S, Sylvain MB, Frederic M, et al. Analysis of hu- man papillomavirus type 16 (HPV16) DNA load and physical state for identification of HPV16-infected women with high-grade lesions or cervical careinoma[J]. J Clin Microbio, 2008,46 ( 11 ) : 3678-3685.
  • 8Akil N, Yasmeen A, Kassab A, et al. High-risk human papil- lomavirus infections in breast cancer in Syrian women and their association with Id-1 expression:a tissue mieroarray study[J]. Br J Cancer, 2008,99 (3) : 404-407.
  • 9Storrs CH, Silverstein SJ. PATJ, a tight j unction-associat- ed PDZ protein is a novel degradation target of high-risk HPV E6 and the alternatively spliced isoform 18 E6[J]. J Virol, 2007,81(8) :4080-4090.
  • 10An,JB,Mo DQ,Liu HR,et al. Inactivation of the CYLD Deubiquitinase by HPV E6 Mediates Hypoxia-Induced NF-κB Activation[J]. Cancer Cell,2008,14(5): 394-407.

引证文献2

重庆医学
2010年 第1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