Establishment of stable SGC-7901 gastric cancer cell line with cortactin down-regulation

Abstract

Objective To establish a stable SGC-7901 gastric cancer cell line with cortactin downregulation.Methods Three pairs of primers specific to knockdown cortactin and a pair of control primers were designed and synthesized.The primers were annealed and inserted into PLKO.1 plasmids.The plasmids,psPAX2,and PCMV-VSVG were transfected into HEK293T cells simultaneously,and then lentiviral particles were produced and collected.The lentiviral particles were used to infect SGC-7901 cells and puromycin was used to select stable cell lines.Three stable cell lines were selected at last.Real-time quantitative polymerase chain reaction (Real-time PCR) and Western blotting were conducted to detect the expression level of cortactin mRNA and protein respectively.Results The expression levels of cortactin mRNA and protein were significantly down-regulated.The mRNA and protein levels of cortactin in PLKO.1 + short hairpin RNA (shRNA)3 cell line were significantly decreased,about 10％ of the control stable cell line.Conclusion The stable SGC-7901 gastric cell lines with cortactin down-regulation were established successfully. Key words: Cortactin; Short hairpin RNA ; Gastric