Effect Of STGC3-siRNA On The Growth In NIH3T3 Cell Line

Objective: STGC3 gene is a candidate tumor suppressor gene cloned recently. NIH3T3 cell line was selected as a target with positive expression of STGC3 gene and observed the proliferation effect after STGC3 gene expression inhibited by siRNA.Methods: pSilencer3.1-STGC3-siRNA expression vector targetting STGC3 gene was constructed and transfected into NIH3T3 cell line by cationic liposome. The stable pSilencer3.1-STGC3-siRNA/NIH3T3 cell line was obtained by G418 selection. The levels of STGC3 mRNA and protein were detected by RT-PCR, western blotting and immunohistochemistry. Cell morphology, growth curves, colony formation rate and cell cycle distribution were detected in NIH3T3 cell line after STGC3 gene expression down-regulated.Results: The results of restriction enzyme cutting and DNA sequencing demonstrate that the pSilencer3.1-STGC3-siRNA expression vector was successfully constructed. After pSilencer3.1-STGC3-siRNA and pSilencer3.1 transfected into NIH3T3 cell, selected with G418 and identified by RT-PCR, western blotting and immunocytochemistry assays, the stable pSilencer3.1-STGC3-siRNA/NIH3T3 cell line of STGC3 gene down-regulated was established successfully. Cell morphology showed that nucleus color became heavier and cellular shape became slimmer in pSilencer3.1-STGC3-siRNA/NIH3T3 cell than the controls.The growth capacity and clone formation potential of pSilencer3.1-STGC3-siRNA/NIH3T3 cell were significantly enhanced after STGC3 gene down-regulated, compared with the controls(P<0.05). FCM analysis indicated that the ratio of S and G2 phrase of pSilencer3.1-STGC3-siRNA/NIH3T3 cells was markedly higher than that of the other two controls(P<0.05). Conclusion:1.The expression vector of pSilencer3.1-STGC3-siRNA was constructed successfully.2.The pSilencer3.1-STGC3-siRNA/NIH3T3 cell line of STGC3 gene expression down-regulated was established successfully.3.The growth of the NIH3T3 cell line increased after STGC3 gene expression down-regulated in the pSilencer3.1-STGC3-siRNA/NIH3T3 cell line.