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目的构建携带RbAp48基因序列的重组腺病毒,检测其对宫颈癌细胞系SiHa肿瘤细胞行为的影响。方法构建穿梭质粒pDC316-RbAp48,与骨架质粒pBHGloxΔE1,3Cre共转染HEK293细胞,获得复制缺陷型重组腺病毒Ad5-RbAp48,Ad5-RbAp48感染人宫颈癌细胞SiHa,采用免疫印迹法检测RbAp48表达,使用WST-8、细胞计数及软琼脂克隆形成实验,研究Ad5-RbAp48介导RbAp48高表达对SiHa细胞增殖与非锚定依赖性生长的影响。结果成功构建并鉴定重组腺病毒Ad5-RbAp48,Ad5-RbAp48感染SiHa细胞导致RbAp48高效表达,并有效抑制SiHa细胞增殖能力与非锚定依赖性生长能力。结论 RbAp48重组腺病毒可介导RbAp48高效表达,具有抑制宫颈癌细胞肿瘤细胞行为的作用。
Objective To construct the recombinant adenovirus carrying RbAp48 gene sequence and test its effect on the behavior of cervical cancer cell line SiHa. Methods The shuttle plasmid pDC316-RbAp48 was constructed and transfected into HEK293 cells with the backbone plasmid pBHGloxΔE1,3Cre to obtain replication-defective recombinant adenovirus Ad5-RbAp48 and Ad5-RbAp48 to infect human cervical cancer cell SiHa. The expression of RbAp48 was detected by Western blotting, WST-8, cell counting and soft agar colony formation assay were used to study the effect of Ad5-RbAp48-mediated RbAp48 overexpression on proliferation and non-anchorage-dependent growth of SiHa cells. Results Recombinant adenovirus Ad5-RbAp48 was successfully constructed and identified. Ad5-RbAp48 infection of SiHa cells resulted in high expression of RbAp48 and effectively inhibited the proliferation and non-anchorage-dependent growth of SiHa cells. Conclusion RbAp48 recombinant adenovirus can induce the high expression of RbAp48 and inhibit the tumor cell behavior of cervical cancer cells.