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[目的]构建能高效表达人mi R-29b的慢病毒载体,研究其对人肺腺癌A549细胞迁移的影响。[方法]PCR扩增mi R-29b前体序列,克隆入慢病毒表达载体,测序鉴定。包装好的慢病毒感染A549细胞,流式细胞仪分选后用荧光定量PCR检测mi R-29b表达水平,划痕实验观察过表达mi R-29b对A549细胞迁移的影响。[结果]测序结果证明成功构建人mi R-29b过表达慢病毒载体。mi R-29b过表达慢病毒感染效率为88.56%,使A549细胞中mi R-29b表达升高2.78倍。与A549细胞相比,稳定过表达mi R-29b的A549-mi R-29b细胞的迁移能力显著性降低。[结论]成功构建mi R-29b过表达慢病毒载体,过表达mi R-29b可抑制A549细胞的迁移,为进一步研究mi RNA对肿瘤转移的调控作用奠定了基础。
[Objective] To construct a lentiviral vector capable of efficiently expressing human mi R-29b and study its effect on the migration of human lung adenocarcinoma A549 cells. [Method] The mi R-29b precursor sequence was amplified by PCR, cloned into the lentiviral vector and sequenced. The packaged lentivirus infected A549 cells. The expression of mi R-29b was detected by fluorescence quantitative PCR after being sorted by flow cytometry. Scratch experiment was performed to observe the effect of mi R-29b overexpression on A549 cell migration. [Results] The sequencing results demonstrated that the human mi R-29b overexpression lentiviral vector was successfully constructed. mi R-29b over-expressed lentivirus infection efficiency of 88.56%, so that mi R-29b expression in A549 cells increased by 2.78 times. Migration ability of A549-mi R-29b cells overexpressing mi R-29b was significantly reduced compared to A549 cells. [Conclusion] The mi R-29b overexpression lentiviral vector was successfully constructed. Overexpression of mi R-29b could inhibit the migration of A549 cells, which lays the foundation for further study of the mi RNAs regulation on tumor metastasis.