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目的构建凋亡因子Caspase-12的短发夹状RNA(siRNA),观察其在体外对鼠粒单系白血病细胞WEHI-3细胞对凋亡的抵抗作用。方法设计并合成一对针对Caspase-12编码基因的siRNA,构建表达此siRNA的重组质粒psiRNA-Caspase-12,转染入WEHI-3细胞。检测胞内Cas- pase-12的mRNA和蛋白的表达。并检测转染后WEHI-3细胞的抗凋亡能力。结果酶切分析和测序证实构建成功,siRNA对Caspase-12的表达均有明显的抑制作用。染后WEHI-3细胞的抗凋亡能力增加。结论构建的重组质粒能有效抑制Caspase-12基因在白血病细胞株WEHI-3中的表达。并对内质网应激性凋亡具有抵抗能力。
Objective To construct short hairpin RNA (siRNA) of apoptotic factor Caspase-12 and observe its resistance to apoptosis in WEHI-3 cells of murine monocytic leukemia cells in vitro. Methods A pair of siRNA encoding Caspase-12 gene was designed and synthesized. The recombinant plasmid psiRNA-Caspase-12 was constructed and transfected into WEHI-3 cells. Detection of intracellular Caspase-12 mRNA and protein expression. The anti-apoptotic ability of WEHI-3 cells after transfection was tested. Results Enzyme digestion analysis and sequencing confirmed the successful construction of siRNA. Caspase-12 expression was significantly inhibited by siRNA. The anti-apoptotic ability of WEHI-3 cells after staining increased. Conclusion The recombinant plasmid can effectively inhibit the expression of Caspase-12 gene in leukemia cell line WEHI-3. And resistance to endoplasmic reticulum stress-induced apoptosis.