人源FOXG1基因真核表达载体的构建、表达及其对HeLa细胞增殖、迁移和侵袭能力的影响  

Construction and expression of human FOXG1 gene eukaryotic expression vector and its effect on HeLa cell proliferation,migration and invasion

作　　者：王雨晴 姜慧杰 陈艳 刘永新 来明名 WANG Yuqing;JIANG Huijie;CHEN Yan;LIU Yongxin;LAI Mingming(School of Basic Medicine,Dali University,Yunnan Dali 671000,China;Shenshan Center of Sun Yat-sen Memorial Hospital,Sun Yat-sen University,Guangdong Guangzhou 510000,China)

机构地区：[1]大理大学基础医学院,云南大理671000 [2]中山大学孙逸仙纪念医院深汕中心,广东广州510000

出　　处：《现代肿瘤医学》2022年第10期1722-1728,共7页Journal of Modern Oncology

基　　金：国家自然科学基金(编号:81660465);云南省应用基础研究计划项目(编号:2014FD045);云南省教育厅科学研究基金研究生项目(编号:2019Y0267)。

摘　　要：目的:构建人源FOXG1真核表达载体,瞬时转染HeLa细胞实现FOXG1过表达,观察FOXG1对HeLa细胞增殖、迁移和侵袭能力的影响,为后续研究人源FOXG1基因在肿瘤中的作用机制奠定理论基础。方法:利用One Step Cloning法构建重组质粒pEGFP-C1-FOXG1,并通过酶切、测序对重组质粒进行鉴定;采用脂质体介导的转染法将pEGFP-C1-FOXG1瞬时转染至HeLa细胞,荧光显微镜下观察绿色荧光蛋白的表达,Western blot检测HeLa细胞中EGFP-FOXG1融合蛋白的表达;进一步,分别通过CCK8、伤口愈合和Transwell实验检测FOXG1对HeLa细胞增殖、迁移和侵袭能力的影响。结果:酶切鉴定和测序表明重组质粒pEGFP-C1-FOXG1构建成功;瞬时转染HeLa细胞24小时后,能够在荧光显微镜观察到绿色荧光,48小时后转染效率达到90%;Western blot显示EGFP-FOXG1融合蛋白正确表达;CCK8实验表明过表达FOXG1能够促进HeLa细胞的增殖;伤口愈合和Transwell实验表明FOXG1过表达可以促进HeLa细胞的迁移和侵袭。结论:成功构建了pEGFP-C1-FOXG1重组质粒,并在HeLa细胞中正确表达;功能实验表明FOXG1能够促进HeLa细胞增殖、迁移和侵袭。Objective:To construct and express human FOXG1 eukaryotic expression vector in HeLa cells,and investigate the effect of FOXG1 on the proliferation,migration and invasion of HeLa cells.Methods:One step cloning method was used to construct the human FOXG1 eukaryotic expression vector.The recombinant plasmid pEGFP-C1-FOXG1 was verified by enzyme digestion and DNA sequencing,and then transient transfected into HeLa cells by liposome.The pEGFP-C1-FOXG1 expression was detected via Western blot.The viability of HeLa cells was measured by CCK-8 assay.Cell wound healing assayand transwell assay were used to study the effects of FOXG1 on the proliferation,migration and invasion of HeLa cells.Results:Enzyme digestion and DNA sequencing confirmed that the pEGFP-C1-FOXG1 vector was successfully constructed.The expression and location of EGFP-FOXG1 fusion protein in the nucleus of HeLa cells could be observed under fluorescence microscopy.Western blot showed that the EGFP-FOXG1 fusion protein was expressed correctly in HeLa cells.CCK-8 assay showed that overexpression of FOXG1 promoted the proliferation of HeLa cells.Cell wound healing assay and transwell assay revealed that FOXG1 could increase the migration ability and the invasion ability of HeLa cells.Conclusion:Eukaryotic expression vector pEGFP-C1-FOXG1 was successfully constructed and expressed in HeLa cells.Functional experiments show that FOXG1 can promote HeLa cell proliferation,migration and invasion levels.

关 键 词：FOXG1 HELA细胞 过表达 增殖 迁移 侵袭 

分 类 号：R730.23[医药卫生—肿瘤]