关键词: siRNA, 人BMP9, 重组腺病毒载体, 肿瘤增殖

Abstract: Objective To screen specific small interfering RNA(siRNA) target human BMP9 gene and prepare recombinant adenovirus vector AdsiBMP9 then investigate its effects on the proliferation of breast cancer SK-BR-3cells. Methods Three pairs of double-stranded DNA fragments for silencing human BMP9 were designed and synthesized, then subcloned into the shuttle plasmid Pses-Hus. The recombinant plasmids Pses-Hus-siBMP9 were transfected into the breast epithelial cells HBL-100 by using lipofectamine transfection reagent, Screened the effective interfering plasmid, constructed AdsiBMP9 and infected SK-BR-3 cells. The expression level of BMP9 mRNA and protein were detected by RT-PCR and western blot. The proliferation of SK-BR-3 cells were observed with MTT assay. Results The recombinant plasmid Pses-Hus-siBMP9 and recombinant adenovirus AdsiBMP9 were successfully constructed and its titer was 1×1010IU/ml. Compared to the negative and non-infected controls, the expression of BMP9 gene was significantly inhibited after the SK-BR-3 cells were infected by AdsiBMP9(P<0.05). SK-BR-3 cells infected with AdsiBMP9 showed a promoted proliferation effect. On the fifth day, the growth rate of experiment groups was significantly higher than that of negative groups(P<0.05). Conclusion Specific siRNA targeting human BMP9 gene was successfully constructed, which can effectively inhibit endogenous expression of BMP9 in SK-BR-3cells and promote its proliferation.

Key words: siRNA, human BMP9, recombinant adenovirus vector, tumor proliferation