利用CRISPR/Cas9系统敲除ITGB6基因对人结肠癌HT-29细胞生物学行为的影响  

Effects of ITGB6 gene knock⁃out by CRISPR/Cas9 on biological behaviors of human HT⁃29 colon cancer cells

作　　者：薛栋 孙祺 邬君义 佘军军[1] 李凡妮 XUE Dong;SUN Qi;WU Junyi;SHE Junjun;LI Fanni(Department of General Surgery,the First Affili-ated Hospital of Xi′an Jiaotong University,Xi′an 710061,China;不详)

机构地区：[1]西安交通大学第一附属医院普通外科,西安710061 [2]西安交通大学第一附属医院人才高地科研平台,西安710061

出　　处：《实用医学杂志》2023年第5期585-590,共6页The Journal of Practical Medicine

基　　金：国家自然科学基金资助项目(编号:81803026,81702362)。

摘　　要：目的 探讨整合素β6(ITGB6)基因敲除后对人结肠癌HT-29细胞生物学行为的影响。方法 通过CRISPR/Cas9技术构建ITGB6敲除的HT-29细胞(ITGB6-KO)作为敲除组,选择空白转染细胞作为对照组。通过Western blot法检测ITGB6表达,通过MTT实验、平板克隆形成实验和Transwell侵袭实验检测敲除ITGB6基因对HT-29细胞的增殖能力、克隆形成能力和侵袭能力的影响。结果 Western blot和DNA测序结果显示CRISPR/Cas9技术有效敲除ITGB6基因。MTT实验、平板克隆形成实验和Transwell侵袭实验显示敲除ITGB6基因能够抑制HT-29细胞增殖能力、克隆形成能力和侵袭能力(P <0.01)。结论 CRISPR/Cas9技术能够有效建立敲除ITGB6基因的结肠癌HT-29细胞系,ITGB6敲除后HT-29细胞恶性生物学行为受到抑制,ITGB6基因有望成为结肠癌治疗的潜在靶点。Objective To investigate the effect of ITGB6 gene knockout on the biological behavior of human HT⁃29 colon cancer cells.Methods We constructed the ITGB6 knock⁃out HT⁃29 cells by CRISPR/Cas9 technolo⁃gy(ITGB6⁃KO),which were defined as the knockout group.Meanwhile,the blank transfected cells were defined as the control group.Western blotting was used to detect the expression of ITGB6 in HT⁃29 cells.MTT assay,Plate colony formation assay and Transwell assay were used to detect the effects of ITGB6 knockout on the ability of proliferation,colony formation and invasion of HT⁃29 cells.Results Western blotting and DNA sequencing showed that CRISPR/Cas9 technology could effectively knock out ITGB6 gene.MTT assay,Plate colony formation assay and Transwell assay confirmed that knocking out ITGB6 gene could significantly inhibit the proliferative ability,colony formation ability and invasive ability of HT⁃29 cells(P<0.01).Conclusion CRISPR/Cas9 technology can effectively establish a ITGB6 knock⁃out HT⁃29 cell line.After knocking out ITGB6 gene,the malignant biologi⁃cal behavior of HT⁃29 cells is inhibited,therefore ITGB6 gene is expected to become a potential target for the treat⁃ment of colon cancer.

关 键 词：结肠癌 ITGB6 CRISPR/Cas9 克隆增殖 侵袭 

分 类 号：R735.3[医药卫生—肿瘤]