Lentivirus-mediated small interfering RNA silencing aryl hydrocarbon receptor nuclear translocator gene in human colon cancer cells HT29

Abstract

Objective To obtain the recombinant lentivirus vector with expressing aryl hydrocarbon receptor nuclear translocator (ARNT)-short hairpin RNA (shRNA) that could suppress the expression of ARNT gene in human colorectal cancer HT29 cells effectively.Methods Construct four lentiviral vector with expressing ARNT-shRNA and a negative control lentiviral vector and have the sequencing appraisal.After the five recombinant lentivirus vector were transfected into the HT29 cells,gain five stable transfection cell lines through enlarging cultivation of cloning dilution,the ARNT mRNA and protein expression were examined by real-time quantitative polymerase chain reaction and Western blotting,respectively.Results Four groups of recombinant lentivirus vector that could express ARNT-shRNA were successfully constructed.One of the groups (LV-ARNT-RNAi-3) could significantly suppress the expression of ARNT gene in HT29 cells,the mRNA expression fell by 70％ (P ＜ 0.05) and the protein expression fell by 60％ (P ＜0.05),respectively,compared with the control group.Conclusion LV-ARNT-RNAi-3 can efficiently inhibit ARNT expression in HT29 cells. Key words: Lentivirus; Aryl hydrocarbon receptor nuclear translocator; RNA interference; HT29 cell line