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采用1个载体编码2个shRNA技术,构建Livin基因和Survivin基因联合靶向的siRNA重组表达载体,研究其对人大肠癌细胞生长协同抑制的增效作用。构建Livin基因和Survivin基因联合靶向的siRNA重组表达载体并转染大肠癌细胞,通过RT-PCR和Western blotting方法检测Livin基因与Survivin基因的表达,利用流式细胞仪检测处理后细胞的凋亡效应。经酶切鉴定和测序分析证实Livin基因和Survivin基因联合靶向的siRNA重组表达载体构建成功,它对大肠癌细胞Livin和Survivin mRNA的抑制率分别为27.90%和32.24%,对Livin和Survivin蛋白表达的抑制率分别为22.28%和40.86%,诱导肿瘤细胞凋亡率为(11.69±1.37)%,但协同干扰作用比单独干扰Livin基因或Survivin基因弱。Livin基因和Survivin基因联合靶向的siRNA重组表达载体构建成功并能抑制Livin基因和Survivin基因的表达,但协同抑制作用比单独干扰Livin基因或Survivin基因弱。
A vector encoding two shRNAs was used to construct a recombinant expression vector targeting Livin gene and Survivin gene to study the synergistic effect of synergistic inhibition on human colorectal cancer cell growth. To construct recombinant siRNA targeting Livin gene and Survivin gene and transfect it into colorectal cancer cells. The expression of Livin gene and Survivin gene was detected by RT-PCR and Western blotting, and the apoptosis of cells was detected by flow cytometry effect. Restriction endonuclease analysis and sequencing analysis confirmed that Livin gene and Survivin gene combined targeting siRNA recombinant expression vector was successfully constructed. The inhibitory rates of Livin and Survivin mRNA in colorectal cancer cells were 27.90% and 32.24%, respectively. The expression of Livin and Survivin protein The inhibitory rates were 22.28% and 40.86%, respectively. The apoptosis rate of induced tumor cells was (11.69 ± 1.37)%, but the synergistic effect was weaker than that of Livin gene or Survivin gene alone. Livin gene and Survivin gene targeting siRNA recombinant expression vector was successfully constructed and can inhibit the expression of Livin gene and Survivin gene, but synergistic inhibition than interfering Livin gene alone or Survivin gene weak.