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Inhibition of GSK-3β Gene Expression by RNA Interference and Its Effect on the Nogo-A Inhibition to Axon Outgrowth of PC12 Cells

Author: DuanXiaoWei
Tutor: ZhaoHongYang
School: Huazhong University of Science and Technology
Course: Neurosurgery
Keywords: Nogo-A GSK-3β RNA interference PC12 cells Axons
CLC: R346
Type: Master's thesis
Year: 2010
Downloads: 49
Quote: 0
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Abstract


Objective To study the GSK-3β gene silencing of Nogo-A inhibit neurite outgrowth of PC12 cells . The constructed targeting GSK-3β short hairpin RNA (shRNA) eukaryotic expression vector , the interference plasmids were transfected using liposomes , blank plasmid and negative control plasmid to PC12 cells , fluorescence inverted microscope 24h after transfection the transfected and reverse transcriptase polymerase chain reaction for detection of the inhibitory effect of GSK-3βRNA . Plus mouse nerve growth factor-induced PC12 differentiation , promote axonal growth after 72 hours to adjust the cell density and add Nogo-A protein observed in the case of the presence of Nogo-A protein fluorescence inverted microscope , 48h , each group PC12 the different cell axon growth changes , using the Image-Pro Plus6.0 software axons of cells measured using SPSS software for statistical analysis . (1) successfully constructed for GSK-3β shRNA eukaryotic expression vector pGenesil1.1 - GSK -3β ; ( 2 ) blue excitation light , successfully transfected cells compared with non-transfected cells can be green fluorescence . The experimental group, the transfection efficiency of the blank control group and negative control group were 43% , 80%, 76% ( 3) interfere with the plasmid can effectively inhibit the gene expression of the GSK-3β ; (4) induced by nerve growth factor in mice in each group cells 72h and 48h after cell axon length of the control group and negative control group had no significant difference ( P gt ; 0.05) ; experimental group cell axons blank group and negative control group growth ( P lt ; 0.05 Nogo-A protein effect ) , there are statistically significant . Conclusion GSK-3β involved in the inhibition of Nogo-A in PC12 cells axon growth .

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