ObjectiveTo investigate the effect of nuclear receptorRev-erbβknockout on proliferation and migration ability of human hepatocellular carcinoma cell line HepG2.Methods-TheRev-erbβgene knockout HepG2 cell line was abtained by CRISPR/Cas9 genome editing technique with specific DNA modification of the target gene. TheRev-erbβgene targeting vectors were co-transfected into HepG2 cells. Through cloning and screening, theRev-erbβgene knockout HepG2 cell line was constructed, PCR, sequencing and Western blot methods were carried out for the identification of theRev-erbβgene knockout HepG2 cell line. The expression level of tumor migration and invasion-associated gene inRev-erbβgene knockout cell was determined by real-time quantitative PCR (qRT-PCR) and was compared with normal cell as control.MTT, cell scratch and Transwell experiments were conducted in order to explore the effect ofRev-erbβgene on HepG2 cell's ability of proliferation, migration and invasion.ResultsARev-erbβgene knockout monoclonal cell line, which was identified by PCR, sequencing and Western blot, was successfully constructed and named HepG2 C5 (Rev-erbβ-/-). qRT-PCR results showed thatRev-erbβknockout resulted in up-regulation of matrix metalloproteinase-2 (MMP2), matrix metalloproteinase-9(MMP9)and extracellular matrix protein-1 (ECM1) gene expression (P< 0.05) and down-regulation of e-cadherin (CDH1) gene expression (P=0.05).Results of MTT, cell scratch and transwell experiments showed that HepG2 C5 had stronger proliferation, migration and invasion ability than control cells (P< 0.05).ConclusionRev-erbβgene knockout could change the expression of migration and adhesion-associated genes in HepG2 cell, and then affect the proliferation, migration and invasion ability of HepG2 cells.
