人类乳头瘤病毒16全基因在体外培养的正常人角质形成细胞 …

目的 利用含有人类乳头瘤病毒（ＨＰＶ）１６全基因的质粒转染正常人角质形成细胞,观察ＨＰＶ１６ｍＲＮＡ在转染细胞的表达。方法 用ＦｕＧＥＮＥ＾ＴＭ６转染试剂,将携带ＨＰＶ１６全基因的质粒ｐＳＶ２－ｎｅｏ／１６转染体外培养的正常人角质形成细胞,在转染后２４ｈ提取细胞总ＲＮＡ和ＤＮＡ,进行ＲＴ－ＰＣＲ和Ｓｏｕｔｈｅｒｎ印迹分析,结果 ２４ｈ后,ＲＴ－ＰＣＲ成功地扩增出１１０ｂｐ的片段,转染细胞中已出现Ｈ

TransientExpression of a Transfected HPV16 Whole Gene in Normal Human Keratinocytes

Objective To investigate the expression of HPV16 mRNA in normal human keratinocytes transfected with a plasmid containing HPV16 whole gene. Methods Normal human keratinocytes were cultured in vitro with HPV16 whole gene and FuGENETM 6, a transfecting reagent, then total RNA and DNA were extracted. RT-PCR and Southern blot were used to identify the expression of HPV16 mRNA and DNA, respectively. Results Tweenty four hours after transfection HPV mRNA was expressed in transfected human keratinocytes and a 110 bp product was successfully amplified by RT-PCR method.A 7.9 kb DNA fragment, which corresponded to HPV16 whole gene, was confirmed in the transfected keratinocytes by Southern blot analysis. Conclusion Tweenty four hours after transfection of normal human keratinocytes with pSV2-neo/16,the expression of HPV16 mRNA and DNA was successfully detected.