Construction of TGF-β2-targeting siRNA interference recombinant plasmids

Abstract:

AIM:To construct specific siRNA expression vector targeting human TGF-β2.METHODS:The mRNA sequences of human TGF-β2 in NCBI were analysed in silico.The best sequence of siRNA were designed according to the siRNA designing principle.The double-stranded DNA was inserted into the GPU6/GFP/Neo vector by the BamH Ⅰ and Bbs Ⅰ.The recombinant plasmid was identified by the BamH Ⅰ and PstⅠ single enzyme digestion and verified by DNA sequencing further.RESULTS:The positions of 1016-1034 in TGF-β2 mRNA were chosen as the RNAi target.The result of the enzyme digestion to the recombinant plasmid showed that the double-stranded DNA was successfully inserted into the GPU6/GFP/Neo vector.The inserted sequence was the same as the designed sequences and was proved by the DNA sequencing.CONCLUSION:The human TGF-β2 specified siRNA interference plasmid was successfully constructed,and this plasmid can be used for the study of the function of TGF-β2 gene and improve the success rate of glaucoma filtration surgery by gene therapy.