摘要
目的研究人tRNA甲基转移酶9样蛋白KIAA1456的高表达对HO8910PM细胞增殖及细胞周期的影响。方法利用PCR扩增KIAA1456编码区片段,克隆插入载体Ubi-MCS-EGFP-IRES-puromycin中,构建Ubi-KIAA1456-EGFP-puromycin(LV-KIAA1456)慢病毒表达载体;将慢病毒表达载体包装生成慢病毒颗粒并感染HO8910PM细胞,72 h后观察绿色荧光蛋白(GFP)在HO8910PM卵巢癌细胞中的表达分布情况;反转录PCR检测HO8910PM细胞中KIAA1456 mRNA水平,Western blot法检测HO8910PM细胞KIAA1456蛋白表达水平;CCK-8法检测细胞增殖活性,流式细胞术检测细胞周期。结果成功构建重组慢病毒表达载体LV-KIAA1456,慢病毒颗粒感染HO8910PM细胞后,细胞中KIAA1456的mRNA和蛋白均高表达。过表达KIAA1456的HO8910 PM细胞增殖能力明显抑制、G1期细胞明显增加。结论过表达KIAA1456显著抑制HO8910 PM卵巢癌细胞增殖,细胞周期阻滞于G1期。
Objective To investigate the effect of over-expressed KIAA1456 on the proliferation and cell cycle of HO8910PM cells. Methods KIAA1456 was amplified by PCR and inserted into lentiviral expression vector Ubi-MCS-EGFP- IRES-puromycin to construct Ubi-KIAA1456-EGFP-puromycin （LV-KIAA1456） vector. Thereafter, the lentiviral particles were packaged and used to infect HO8910PM cells. The green fluorescence protein （GFP） expression was observed at 72 hours. The expression of KIAAI456 was examined by reverse transcription PCR and Western blotting. Cell growth was detected with CCK-8 assay and cell cycles were analyzed by flow cytometry. Results Reverse transcription PCR and Western blotting showed that HO8910PM cells infected with LV-KIAA1456 vector over-expressed KIAA1456 mRNA and protein. Cell proliferation was inhibited and G1 phase cells significantly increased in HO8910PM cells overexpressing KIAA1456. Conclusion Over- expression of KIAA1456 inhibits the proliferation of HO8910PM ovarian cancer cells and arrests the cell cycle in G1 phase.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2016年第9期1183-1187,共5页
Chinese Journal of Cellular and Molecular Immunology
基金
重庆市渝中区科技计划(20150116)
重庆市妇科微创诊治特色专科项目(渝卫科教2013[46]号)
