生物学杂志

生物学杂志 ›› 2022, Vol. 39 ›› Issue (1): 11-.doi: 10. 3969/ j. issn. 2095-1736. 2022. 01. 011

• 研究报告 • 上一篇    下一篇

慢病毒介导SHCBP1 低表达对黑色素瘤B16细胞增殖的影响

  

  1. 陕西理工大学 生物科学与工程学院,汉中723000
  • 出版日期:2022-02-18 发布日期:2022-02-15
  • 通讯作者: 路宏朝,博士,教授,研究方向为肿瘤发生分子机制研究,E-mail:zl780823@ 126. com
  • 作者简介:孙志阳,硕士研究生,研究方向为肿瘤发生分子机制研究,E-mail:460084857@ qq. com
  • 基金资助:
    陕西省教育厅科技计划项目(No. 20JK0570)

The effect of lentivirus-mediated down-regulation of SHCBP1 on the proliferation of melanoma B16 cells#br#

  1. The effect of lentivirus－mediated down－regulation of SHCBP1on the proliferation of melanoma B16 cells
  • Online:2022-02-18 Published:2022-02-15

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摘要： 构建Shcbp1 慢病毒干扰载体,探究SHCBP1 在黑色素瘤B16 细胞中的生物学功能。通过设计靶向干扰小鼠Shcbp1 基因的shRNA-1 和shRNA-2 以及对照序列shRNA-NC,连接至慢病毒骨架载体,利用HEK293T 细胞包装慢病毒颗粒,侵染B16 细胞,检测干扰效率,并通过CCK-8 和流式细胞术检测细胞活力和细胞周期。利用WesternBlot 检测增殖相关基因的表达。结果显示:敲低Shcbp1 显著抑制B16 细胞增殖;流式细胞术检测结果发现,敲低Shcbp1 将B16 细胞阻滞在 G1 期;ERK1/2 的磷酸化水平降低,P21 的表达水平升高。结果表明SHCBP1 可能通过ERK1/2 信号通路抑制P21 的转录,加速B16 细胞从G1 到S 期进程。

关键词: SHCBP1, 细胞增殖, B16 细胞, 慢病毒干扰

Abstract: To construct Shcbp1 lentiviral interference vector to explore the biological function of SHCBP1 in melanoma B16 cells, first,the shRNA-1 and shRNA-2 that target to interfere with the mouse Shcbp1 gene and the control sequence shRNA-NC were designed and connected to the lentiviral backbone vector, HEK293 cells were used to package the lentiviral particles to infect B16 cells, the interference efficiency was tested and CCK-8 and flow cytometry were used to detect cell viability and cell cycle. Western Blot was used to detect the expression of proliferation-related genes. The results showed that knockdown of Shcbp1 significantly inhibited the proliferation of B16 cells; flow cytometry results showed that knockdown of Shcbp1 blocked B16 cells in the G1 phase; the phosphorylation level of ERK1/2 decreased and the expression level of p21 increased. These results indicate that SHCBP1 may inhibit the transcription of P21 through the ERK1/2 signaling pathway and accelerate the progress of B16 cells from G1 to S phase

Key words: Shcbp1, cell proliferation, B16 cell, lentiviral interference

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