siRNA对体外培养大鼠海马神经元细胞红蛋白基因表达的抑制

发布时间：2018-08-04 10:21

【摘要】：【目的】研究siRNA(small interfering RNA)对细胞红蛋白(Cytoglobin,CYGB)基因表达的抑制作用。【方法】取新生大鼠海马做神经元原代培养,NeuN免疫荧光抗体鉴定神经元纯度后,通过阳离子脂质体转染试剂将体外化学合成的针对CYGB基因的siRNA转入细胞中,以未转入siRNA细胞及转入阴性siRNA细胞为对照,通过转染荧光对照siRNA检测转染效率,用RT-PCR方法检测siRNA对CYGB表达的抑制作用。【结果】成功体外培养海马神经元,转染荧光对照siRNA结果示转染效率在70%左右,RT-PCR结果提示与阴性对照及空白对照相比,转染siRNA-78后24 h(P0.01),48 h(P0.001)及60 h(P0.001)CYGB mRNA的表达明显减弱,72 h后表达无明显减少。而转染siR-NA-79,siRNA-80在各时间点与对照相比没有明显差别(P0.05)。【结论】特异性针对CYGB的siRNA-78转染体外培养的大鼠海马神经元24~60 h能显著下调CYGB基因的表达。
[Abstract]:[objective] to study the inhibitory effect of siRNA (small interfering RNA) on the expression of Cyb gene. [methods] Neun immunofluorescence antibody was used to identify the purity of the neurons from hippocampal neurons of newborn rats. The chemically synthesized siRNA targeting CYGB gene was transferred into the cells by cationic liposome transfection reagents. The transfection efficiency was detected by transfection of siRNA, which was not transfected into siRNA cells and negative siRNA cells. RT-PCR method was used to detect the inhibitory effect of siRNA on the expression of CYGB. [results] Hippocampal neurons were successfully cultured in vitro. The transfection efficiency of fluorescent control siRNA was about 70%. The results of RT-PCR showed that compared with negative control and blank control, the transfection efficiency was about 70%. 24 h (P0.01) 48 h (P0. 001) and 60 h (P0. 001) after siRNA-78 transfection, the expression of P0. 001) CYGB mRNA decreased significantly, and the expression of P0. 001) CYGB mRNA did not decrease after 72 h. However, there was no significant difference between siR-NA-79 siRNA-80 and control at each time point (P0.05). [conclusion] the expression of CYGB gene was significantly down-regulated in rat hippocampal neurons transfected with CYGB specific siRNA-78 for 2460 h.
【作者单位】：汕头大学医学院第二附属医院儿科;
【基金】：广东省科技计划项目(2008B080701036) 广东省医学科研基金立项(B2009180)
【分类号】：Q343

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