Construction of siRNA Specific for Smad2 and Identification of its Efficiency in Carcinoma Cell Line

Objective To construct the small interfering RNA(siRNA) eukaryotic expression vector specific for smad2 and to observe its silencing effect on smad2 gene in carcinoma cells.Methods Oligonucleotides were designed specific for smad2 gene and cloned into siRNAs expression vector Psilencer2.1-U6 neo,then transformed into E.coli DH5α.The expression vectors were identified by enzyme digestion and sequence analysis.The smad2 siRNA vector was transfected or co-transfected with Flag-smads expression vectors into 293T cells and Hela cells.The expression of smad2 was analyzed by Western blot.Results The result of restriction enzyme digestion and nucleotide sequencing confirmed that the recombinant plasmids were correct.In transfected 293T cells and HeLa cells,Smad2 protein expression level was suppressed over 70%.Conclusion The Psilencer2.1-U6/siRNA specific for smad2 were successfully constructed,which laid foundation for further study of smad2 in base research of cancer.