南方医科大学学报

南方医科大学学报 ›› 2016, Vol. 36 ›› Issue (03): 375-.

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TALEN介导的MYH9 基因沉默及对细胞周期与凋亡的影响

朱显军，邓海军，叶耿泰，沈智勇，李风萍，郭伟洪，杨庆斌，刘浩，李国新   

  • 出版日期:2016-03-20 发布日期:2016-03-20

TALEN-mediated MYH9 Knock-down and its influence on cell cycle and apoptosis of MGC803 cell line

  • Online:2016-03-20 Published:2016-03-20

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摘要： 目的利用TALEN技术敲除人胃癌细胞系MGC803细胞株MYH9基因，观察MYH9基因沉默后细胞周期及凋亡改变。 方法根据斯丹赛FastTALETM TALEN试剂盒说明书，设计并构建靶向MYH9基因的TALEN质粒对。通过质粒转染、DNA测 序、RT-PCR和Western blot等检测质粒活性，成功挑取MYH9基因敲低单克隆株，并对构建好的细胞株进行周期和凋亡检测。 结果成功挑选的MGC803单克隆细胞株未检测到MYH9基因完全敲除；MYH9基因敲低后，MGC803细胞周期受阻于G2/M 期（P<0.05），早期凋亡增加（P<0.05）。结论利用TALEN技术成功构建MGC803细胞MYH9基因敲低单克隆株，该模型有助于 后期深入探讨胃癌MYH9基因功能。

Abstract: Objective To construct a MYH9 gene knockout model in MGC803 cell line using transcription activator-like effector nuclease (TALEN) and observe its effect on cell cycle and apoptosis. Methods According to FastTALETM TALEN Kit, we designed TALEN pairs and constructed the plasmids targeting to MYH9 gene. After detecting their activity in MGC803 cells by plasmid transfection, DNA sequencing, RT-PCR and western blot, we selected the monoclonal cells and studied the changes in the cell cycle and apoptosis. Results MYH9 gene could not be knocked out but knocked down in selected MGC803 monoclonal cells, which caused cell cycle arrested at G2/M phase (P<0. 05) and a significant increase in the cell number with early apoptosis (P<0. 01). Conclusion We successfully generated a MYH9 knockdown model in MGC803 cell lines by TALEN, which could be in favor of MYH9 function study in gastric cancer.