宫颈癌组织中人乳头瘤病毒16型E7基因的DNA改组

DNA Shuffling of HPV16 E7 gene in Cervical Cancer

目的利用DNA改组技术进化人乳头瘤病毒16型E7(HPV16E7)基因,建立HPV16E7基因突变体库。方法从宫颈癌标本中扩增出HPV16E7基因,用脱氧核糖核酸酶(DNaseⅠ)碎片化该基因,切割成50 bp左右的小片段。这些小片段在不外加引物的条件下,利用PCR反应进行重聚,再将重聚物经过一轮普通PCR扩增。结果电泳切胶回收获得与原片段大小相当的基因片段,改组结果较为成功。结论DNA改组在HPV16E7改组中的应用并成功,有利于从HPV16E7基因突变题库中筛选出高免疫原性的基因型别,并为进一步构建HPV治疗性疫苗奠定基础。

Objective To utilize DNA shuffling for directed evolution of HPV16 E7 gene, and to construct the mutant library of HPV16 E7 gene. Methods HPV16 E7 gene was amplified by PCR from the samples of uterine cervix cancer and digested with DNase Ⅰ . Then the random fragments about 50 bp were purified and reassembled by PCR without primers, and the products were amplified by routine PCR. Results These 50-bp fragments were successfully reassembled to genes of the same size as HPV16 E7. Conclusion The successful DNA shuffling of HPV16 E7 gene in uterine cervical cance may help us screen the high immunogenicity of HPV16 E7 gene,which provides a basis for constructing the therapeutic vaccine for human papillomavirus.