Validation of Sn and CD163 receptor-targeted amiRNAs. (A) The schematic structure of amiRNA expression vector. PCMV, immediate early promoter of cytomegalovirus; miR, miR-flanking sequences of mouse BIC non-coding mRNA; Pre-amiRNA, double–stranded oligonucleotide for amiRNA; TK pA, TK gene poly(A) signal of human simplex herpes virus. (B) The schematic structure of the reporter vectors for amiRNA validation. PCMV, immediate early promoter of cytomegalovirus; Sn/CD163, porcine Sn or CD163 receptor cDNA; GFP, green fluorescent protein coding sequence; SV40 pA, poly(A) signal of SV40 virus. (C or D) NIH 3 T3 cells were transfected with different amiRNA expression vector plus the reporter vector, and the GFP-positive cell numbers were measured by flow cytometry 24 h after transfection. The know-down efficiency of each amiRNA was expressed as the percent inhibition of total fluorescence in the cell culture co-transfected with the amiRNA expression vector and the reporter vector.