The SKOV3 Cell MRNA And Protein Expression Levels Of Beclin1 In Gene Intervention After The Initial Study

ObjectiveTo intervent the expression volume(super or reduction expression) of Beclinl gene in SKOV3cells,detected the differential expression mRNA and proteins in SKOV3by two-dimensional difference gel electrophoresis-mass spectrometry techniques and Gene chip technology.MethodCultivated the ovarian cancer cells SKOV3in vitro, ranked them according to different factors.SKOV3-Beclinl:Beclinl gene super-expression in SKOV3cells(transfection of recombinant vector pcDNA3.1/Beclinl); SKOV3-pcDNA3.1: the control group of the SKOV3-Beclin1group (transfection of pcDNA3.1); SKOV3-siRNA:Beclinl gene ischo-expression in SKOV3cells(transfection of siRNA); SKOV3-RNAiMAX: the control group of the SKOV3-siRNA group(transfection of RNAiMAX); group of control:undisposed SKOV3cells. Test the expression volume of Beclin1gene in the five group by Real-time fluorescence quantitative PCR. Then detected the differential expression mRNA and proteins mRNA between the group of SKOV3-Beclinl, SKOV3-siRNA and SKOV3by two-dimensional difference gel electrophoresis-mass spectrometry techniques and Gene chip technology.Result1, After siRNA feeler of Beclinl gene, super-expression carrier transfected the ovarian cancer cells SKOV3, the outcome of Real-time fluorescence quantitative PCR was that: relative copy volume of Beclinl gene increased significantly in SKOV3transfected which group of SKOV3-Beclinl discoved, the outcome was contradictory in group of SKOV3-siRNA, to explain that we implement the different expression of Beclinl gene in SKOV3successfully.2, We detected by gene chip technology that5110mRNA has changes in expression level,18of them altered accordingly in protein level. They arePTPRK, AMIG02, PDIA3, SCAMCl, MSN, SAG, ANXA3, ANXA5, ERP29, GSTP1, PCK2, MVP, PRDX4, TDPX, BECN1, SOD2, STX5, ZNF929.Detected32differential expression proteins by fluorescent two-dimensional difference gel electrophoresis-mass spectrometry, after SKOV3was interfered in. They are Aldehyde dehydrogenase, mitochondrial, Amphoterin-induced protein2, Annexin A3, Annexin A5, Calcium-binding mitochondrial carrier protein SCaMC-1,Dedicator of cytokinesis protein, Endoplasmic reticulum protein ERp29, Enoyl-CoA hydratase, mitochondrial, Glutaminase kidney isoform, mitochondrial, Glutathione S-transferase P, Hypoxia up-regulated protein1, Major vault protein, Methylmalonyl-CoA epimerase, mitochondrial, Moesin, Myosin-VIIb, NADHdehydrogenase iron-sulfur protein3, mitochondria, NADH-ubiquinone oxidoreductase subunit,mitochondrial, Peroxiredoxin-4, Phosphoenolpyruvate carboxykinase [GTP],mitochondrial, Protein disulfide-isomerase A3, Pre-mRNA-processing factor19,Receptor-type tyrosine-protein phosphatase k,Retinitis pigmentosa1-like1protein S-arrestin, Superoxide dismutase [Mn], mitochondrial, Syntaxin-5Thioredoxin-dependent peroxide reductase, mitochondrial,Zinc finger protein292.Conclusion1, Screened5110genes which altered in mRNA level;32proteins had changes in expression level,18of them altered accordingly in protein level. Receptor-typetyrosine-proteinphosphatase k, Amphoterin-induced protein2, Protein disulfide-isomerase A3, Calcium-binding-mitochondrial-carrier-protein, Moesin, S-arrestin, Annexin A3, Annexin A5, Endoplasmic reticulum protein, Glutathione S-transferase P,GTP,mitochondrial, Major vault protein, Peroxiredoxin-4, Thioredoxin-dependent-peroxide-reductase,Mitochondrial, Beclin1, Superoxide dismutase [Mn], mitochondrial, Syntaxin-5, Zinc finger protein292.