【摘要】 目的:探讨肽核酸(peptide nucleic acid,PNA)阻断PYR复合体(polypyrimidine complex)与DNA序列结合对γ-珠蛋白基因表达的影响。方法:设计并合成PYR-PNA、β-PNA和RS-PNA(随机序列PNA),以阳离子脂质体lipofectamine 2000为载体,将PNA转染到K562细胞中。用RT-PCR和Western blot技术分别在转录水平和蛋白翻译水平检测K562细胞在转染24、48和72 h后γ-珠蛋白基因的表达情况。结果:PYR-PNA组在转染24、48和72 h后γ-珠蛋白基因mRNA和蛋白表达水平均显著高于RS-PNA组和空白对照组(P<0.05),其中以转染48 h后的表达上调最为明显,分别为空白对照组的2.0和2.5倍。结论:PYR-PNA可显著上调K562细胞中γ-珠蛋白基因的表达;本研究为β-地中海贫血基因治疗提供了新的研究思路。更多还原

【Abstract】 Objective: To investigate the effect of blocking polypyrimidine complex binding to DNA site by using peptide nucleic acid( PNA) on γ-globin gene expression.Methods: PYR-PNA,β-PNA and RS-PNA( random sequence-PNA) were designed and synthesized,then were transfected into K562 cells with the cationic liposome lipofectamine 2000 used as vector.The expression of γ-globin gene at both the transcriptional and translational level was detected by RT-PCR and the Western blot respectively at 24 h,48 h and 72 h after transfection with PNAs.Results:Compared with RS-PNA and control groups,the expression of γ-globin gene at mRNA and protein levels in PYR-PNA group was significantly up-regulated( P < 0.05),especially at 48 h after tranfection,the levels of mRNA and protein in PYR-PNA group were increased by 2.0 and 2.5 times than those in control group,respectively.Conclusion: PYR-PNA can significantly up-regulate the expression of γ-globin gene in K562 cells,this study may provide a new research idea for gene therapy of β-thalassemia.更多还原