An optimized recombinant HPV16 E6E7 fusion gene (HPV16 ofE6E7) was constructed according to codon usage for mammalian cell expression, and a mutant of HPV16 ofE6E7 fusion gene (HPV16 omfE6E7) was generated by site-dir...An optimized recombinant HPV16 E6E7 fusion gene (HPV16 ofE6E7) was constructed according to codon usage for mammalian cell expression, and a mutant of HPV16 ofE6E7 fusion gene (HPV16 omfE6E7) was generated by site-directed mutagenesis at L57G, C113R for the E6 protein and C24G, E26G for the E7 protein for HPV16 ofE6E7 [patent pending (CN 101100672)]. The HPV16 omfE6E7 gene constructed in this work not only lost the transformation capability to NIH 3T3 cells and tumorigenicity in SCID mice, but also maintained very good stability and antigenicity. These results suggests that the HPV16 omfE6E7 gene should undergo further study for application as a safe antigen-specific therapeutic vaccine for HPV16-associated tumors.展开更多
Objective: To investigate the expression of HPV16 mRNA innormal human keratinocytes transfected with pSV2-neo/16. Methods: First human keratinocytes were cultured in theserum-free medium M154.Second, the plasmid pSV2-...Objective: To investigate the expression of HPV16 mRNA innormal human keratinocytes transfected with pSV2-neo/16. Methods: First human keratinocytes were cultured in theserum-free medium M154.Second, the plasmid pSV2-neo/16was transfected into the human keratinocytes using atransfecting reagent. Third, RT-PCR and Southern Blottingwere used to detect the expression of HPV16 mRNA and DNAin the transfected keratinocytes, respectively. Results: The expression of HPV 16 mRNA was successfullyamplified and an 110bp was detected by RT-PCR. A 7.9kbfragment was confirmed in the transfected keratinocytes bySouthern Blot analysis. Conclusion: HPV 16 mRNA and DNA were successfullydetected in the human keratinocytes.展开更多
基金National 863 Program(2007AA021107)Beijing Municipal Commission of Education Research Projects(KM201010005008)Doctoral Start-up Foundation of Beijing University of Technology
文摘An optimized recombinant HPV16 E6E7 fusion gene (HPV16 ofE6E7) was constructed according to codon usage for mammalian cell expression, and a mutant of HPV16 ofE6E7 fusion gene (HPV16 omfE6E7) was generated by site-directed mutagenesis at L57G, C113R for the E6 protein and C24G, E26G for the E7 protein for HPV16 ofE6E7 [patent pending (CN 101100672)]. The HPV16 omfE6E7 gene constructed in this work not only lost the transformation capability to NIH 3T3 cells and tumorigenicity in SCID mice, but also maintained very good stability and antigenicity. These results suggests that the HPV16 omfE6E7 gene should undergo further study for application as a safe antigen-specific therapeutic vaccine for HPV16-associated tumors.
文摘Objective: To investigate the expression of HPV16 mRNA innormal human keratinocytes transfected with pSV2-neo/16. Methods: First human keratinocytes were cultured in theserum-free medium M154.Second, the plasmid pSV2-neo/16was transfected into the human keratinocytes using atransfecting reagent. Third, RT-PCR and Southern Blottingwere used to detect the expression of HPV16 mRNA and DNAin the transfected keratinocytes, respectively. Results: The expression of HPV 16 mRNA was successfullyamplified and an 110bp was detected by RT-PCR. A 7.9kbfragment was confirmed in the transfected keratinocytes bySouthern Blot analysis. Conclusion: HPV 16 mRNA and DNA were successfullydetected in the human keratinocytes.
