携带Bcl-xl基因的重组慢病毒的制备和鉴定

Construction and identification of a recombinant lentivirus containing Bcl-xl

目的制备携带Bcl-xl基因的重组慢病毒,并鉴定其感染细胞有效性。方法从本所以前构建的重组质粒pAAV-Bcl-xl获得Bcl-xl基因的编码序列,将其置换掉慢病毒质粒pSin-EF2-SOX2-Pur中的SOX2基因编码序列,从而得到重组慢病毒质粒pSin-EF2-Bcl-xl-Pur。然后利用这一质粒包装制备了携带Bcl-xl基因的重组慢病毒颗粒,用其感染细胞后用Western blot和流式检测仪分析了感染前后细胞中Bcl-xl基因的表达情况。结果感染后细胞中Bcl-xl蛋白的表达水平显著高于感染前。结论携带Bcl-xl基因的重组慢病毒颗粒包装成功,能高效感染细胞,为后续的遗传改造树突细胞打下了基础。

Objective To construct a recombinant lentivirus containing Bcl-xl gene,and to detect its infection capability. Methods Bcl-xl-encoded sequence was obtained from our previously constructed plasmid pAAV-Bcl-xl,and used to substitute the SOX2 sequence in a lentiviral plasmid pSin-EF2-SOX2-Pur to construct a lentiviral plasmid pSin-EF2-Bcl-xl-Pur. The recombinant lentivirus containing Bcl-xl was assembled,and used to infect 293FT cells. Western blot and FACS analysis were used to detect Bcl-xl expression in infected cells. Results The results indicate that the Bcl-xl expression level in the infected cells are significantly higher than that in the control cells. Conclusion The recombinant lentivirus containing Bcl-xl was successfully constructed,which laid a foundation for using Bcl-xl to genetically modify dendritic cells.