关键词: [关键词] OPN, siRNA, HEK-297T, 慢病毒载体

Abstract: Objective To construct the recombinant lentivirus vector and explore its effects on colon cancer cell line SW480. Methods The siRNA interfering sequence targeting to osteopontin gene was designed and synthesized,then DNA segment was gained through annealing after chemosynthesis,and then was cloned to pSicoR vector, the recombinant lentivirus vector was constructed and used to challenge SW480 cells.The expression level of osteopontin mRNA and protein was detected by Real Time PCR and Western blotting. The effects on SW480 cells proliferation were analyzed by MTT assay. Results The restriction enzyme digestion,DNA sequencing and detection of GFP expression demonstrated that recombinant lentivirus vector was constructed successfully. Real Time PCR and Western blot analysis confirmed that the expression of OPN mRNA and protein were inhibited.The MTT assay showed that the anti-proliferation effect of lentivirus vector of osteopontin-siRNA on SW480 cells were significant(P<0.05). Conclusions The recombinant lentivirus vector could effectively inhibit the expression of osteopontin in SW480 cells and depress the proliferation of SW480 cells.

Key words: [Key words] osteopontin, siRNA, HEK-293T, lentivirus vector