Construction and identification of L-arginine transporter CAT-2 siRNA recombinant plasmid

Objective: To construct and identificate the recombinant plasmid expression vector of cationic amino acid transporter-2 siRNA,and establish a foundation to study the influence of target siRNA on the expression of CAT-2 and L-arginine/iNOS/NO pathway.Methods: According to the mRNA sequences of L-arginine transporter CAT-2,siRNA design principles and BLAST Comparison,four pairs single-strand DNA of siRNA were designed and synthesized.Annealing into double-strand DNA,the four pairs of double-strand DNA of the siRNA were inserted into siRNA expression vector pcDNATM 6.2-GW/EmGFPmiR to construct siRNA expression plasmid with vector construction kit BLOCK-iTTM Pol II miR RNAi Expression Vector Kit with EmGFP,and transfected the recombinant plasmid into the competent Escherichia coli DH5.After selecting positive Escherichia coli DH5,and extracting the recombinant plasmids,the sequences of the recombinant plasmids were finally identificated.Results: The sequencing results of the recombinant plasmid expressing CAT-2 siRNA confirmed construction successfully.Conclusion: The recombinant plasmids vectors expressing L-arginine transporter CAT-2 siRNA are successfully constructed,which establish a foundation to study the target unfluence siRNA on the expression of CAT-2 and L-arginine/iNOS/NO pathway.