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目的筛选有效抑制磷酸甘油酸激酶1(PGK1)基因的短发夹核糖核酸(shRNA)序列。方法针对PGK1的不同基因区域设计4个shRNA序列,应用LipofectamineTM2000方法转染至胶质瘤细胞株U251细胞,荧光显微镜下观察shRNA的转染效率。采用RT-PCR方法检测PGK1的mRNA表达,Western blot方法检测PGK1的蛋白表达。结果针对PGK1-homo-441位点的shRNA4(5′-GCAAGGATGTTCTGTTCTTGA-3′)能更有效地抑制PGK1的mRNA及蛋白表达。结论shRNA4是能有效抑制PGK1表达的序列,可用于PGK1的功能研究。
Objective To screen short hairpin RNA (shRNA) sequences that effectively inhibit phosphoglycerate kinase 1 (PGK1) gene. Methods Four shRNA sequences targeting different gene regions of PGK1 were designed and transfected into glioma cell line U251 by LipofectamineTM2000. The transfection efficiency of shRNA was observed under a fluorescence microscope. The mRNA expression of PGK1 was detected by RT-PCR and the protein expression of PGK1 was detected by Western blot. Results shRNA4 (5’-GCAAGGATGTTCTGTTCTTGA-3 ’) directed against the PGK1-homo-441 site inhibited the PGK1 mRNA and protein expression more effectively. Conclusion shRNA4 can effectively inhibit the expression of PGK1 and can be used for PGK1 functional studies.