乳头瘤病毒11型完整基因组成功转染角质形成细胞并促其增殖

Transfecting Keratinocyte and Enhancing Cell Proliferation by Integrated HPV11 DNA

摘要:

构建含人乳头瘤病毒11型(Human papillomavirus type 11, HPV11)完整开放读码框(Open reading frame, ORF)基因组的质粒并对其功能进行初步验证, 为构建HPV11转基因动物模型奠定基础。分别构建重组质粒pQE-Trisystem- EGFP/HPV11(pE/H)、pQE-Trisystem-EGFP/1.1copyHPV11(pE/1.1H), 将pE/1.1H、pE/H、闭环状HPV11基因组分别转染原代人角质形成细胞(Keratinocyte, KC)并进行检测。在质粒上成功构建了目的序列, 转染后在pE/1.1H组、pE/H组、HPV11组中检测到HPV11E6基因的表达; 在pE/1.1H组、pE/H组中检测到荧光; HPV11组、pE/H组、pE/1.1H组具备促细胞增殖功能, 实验组间比较pE/1.1H组稍弱(P<0.01), 但与对照组相比均差异极显著(P<0.01)。将具有完整ORF的HPV11基因组(1.1copyHPV11)成功构建于质粒上, 其具备野生型HPV11病理表型; 为研究低危型HPV致病机理提供了实验材料和方法学指导; 为进一步构建HPV11转基因小鼠奠定了基础。

Abstract:

We constructed the plasmid containing the integrated open reading frame (ORF) HPV11 genome, and identified its function, to lay foundation for production of transgenic animal models of HPV11. Recombinant plasmid pQE-Trisystem- EGFP/HPV11 (pE/H) and pQE-Trisystem-EGFP/1.1 copy HPV11 (pE/1.1H) were constructed. Then, the human keratinocyte (KC) was transfected by pE/1.1H, pE/H and closed circular HPV11 and detected. The recombinant plasmid was successfully constructed. The expression of HPV11E6 gene was detected in the experimental group. Fluorescence was observed in the pE/1.1H and pE/H group. The HPV11, pE/H, and pE/1.1 enhanced the KC proliferation, with remarkable differences (P<0.01) from the control group. Amongst the three experimental groups, pE/1.1H was found to be the weakest. The plasmid containing the integrated ORF of HPV11 (1.1 copy HPV11) genome was successfully constructed. The pE/1.1H had the same phenotype of wild-type HPV11 genome. It provided experimental material and methodological guidance for studying the low-risk HPV, as well as for the production of HPV11 transgenic mice.