Vector construction and silencing effect of VEGF gene targeted small interfering RNA
AIM:To construct VEGF gene-targeted small interfering RNA(siRNA)and its expression vector;and to observe the expression level of its expression vector VEGF gene in human osteosarcoma MG-63 cell transfected by the vector.METHODS:First to design VEGF gene-targeted hairpin siRNA,then to synthesize two complementary oligo nucleotide strand,after annealing,to insert the two-strand oligo nucleotide into pSuper neo vetor,which was then restrictive enzyme digested and sequenced.After that we transfected human osteosarcoma MG-63 cell line with the vector using lipofectamine method.Finally,detected the mRNA expression level of VEGF gene through RTPCR.RESULTS:Restrictive enzyme digestion and sequencing confirmed the vector containing siRNA was what we wanted;RTPCR showed the expression level of VEGF gene in transfected MG-63 was decreased.CONCLUSION:We had constructed the correct VEGF gene-targeted siRNA and its vector,and the vector could obviously reduce the VEGF gene expression after transfecting.
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