Effect Of Silencing EGFR Gene Expression By RNA Interference On The Invasion Of Ovarian Cancer Cells

Objective To construct expression vector of epidermal growth factor receptor(EGFR) specific small interfering RNA(siRNA) and stably transfect it into human ovarian cancer cell line SKOV3.To investigate the effect of RNA interference(RNAi) on the EGFR gene expression,cell growth and the invasion of the cell line.Methods According to the siRNA design principle,we designed specific siRNA targeted EGFR and synthesized it in vitro,and it was directed cloned into pSilencer^TM2,1-U6 neo plasmid.DNA sequencing confirmed whether the insertion sequence was correct or not. Then transfected the recombinant plasmid into human ovarian cancer cell line SKOV3 with lipofection.The positive clones were selected by G418.There were three groups: normal control,non-specific transfect and specific transfect.The inhibitive effect of siRNA on EGFR mRNA and protein level were respe ctively detected by reverse transcription polymerase chain reaction(RT-PCR) and immunocytochemistry.The proliferation of SKOV3 cells was detected by colony formation assay.The invasiveness of SKOV3 cells was detected by Transwell invasion assay.Results DNA sequencing confirmed that the insertion sequence of expression vector of EGFR specific siRNA was completely correct.and it obviously down-regulated the expression of EGFR mRNA(F =50.45,q = 12.87,11.72,P<0.01) and protein(F = 61.54, q = 14.58,11.46,P<0.01),The proliferation of EGFR specific siRNA transfect cell was significantly slower than the other two groups(F=130.12,q=21.12,18.15 P <0.01).The invasion ability was obviously decreased(F=34.09,q=11.26,8.34,P < 0.01).Conclusion The EGFR specific siRNA expression vector can effectively suppress the expression level of EGFR gene of SKOV3 cells and lead to the suppression of proliferation and invasion of SKOV3 cells.