摘要
目的 克隆小鼠肥大细胞瘤P815细胞株的P1A基因以制备肿瘤DNA疫苗。方法 用RT PCR方法制备P1A基因 ,以哺乳细胞高效表达质粒pCI neo为载体 ,构建重组DNA疫苗。重组体用克隆位点上游和下游的T7和T3启动子序列为测序引物 ,用自动测序仪测序鉴定克隆的正确性。再将鉴定过的重组质粒用磷酸钙法转化 2 93细胞 ,用RT PCR法鉴定转化细胞中P1A基因的表达。结果 正确构建了P1A/pCI neo重组质粒 ,并且在转化此质粒的 2 93细胞中检测出了P1A的表达。结论 成功地构建了重组P1A/pCI neo肿瘤疫苗 。
Objective To clone P1A gene from mouse mastocytoma cell strain P815 for the production of tumor DNA vaccine. Methods P1A gene was prepared with RT PCR and then ligated with the vector pCI neo which having high efficient expression in mammals to construct a recombinant DNA vaccine. The recombinant was sequenced by automatic sequencer with promoters T7 in upstream and T3 in downstream as sequencing primers. After the identified recombinant plasmids were transformed into cell 293 with Ca 3(PO 4) 2, the expressions of P1A in the cells were determined with RT PCR.Results The recombinant P1A/pCI neo plasmid was constructed successfully and the expression of P1A gene could be detected in the transformed 293 cells. Conclusion Our experiments indicate that the recombinant P1A/pCI neo tumor DNA vaccine can be constructed successfully which could be used for the studies of tumor animal model.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2000年第10期949-953,共5页
Journal of Third Military Medical University
基金
国家自然科学基金资助项目 !(3970 0 1 32 )
