RNA interfering (RNAi) is a process of sequence-specific, posttranscriptional gene silencing in animals and plants. In mammalian cells, duplexes of 19～25 nts (nucleotides) RNAs efficiently inhibit gene expression. The pBS/H1SP vector expressing siRNA is used which inhibit specific HIV-1 gene expression. To assess the intracellular effect of this H1 promoter-driven siRNA, a reporter plasmid pEGFP-C1-HIV protein which expresses fusion protein of enhanced green fluorescent protein (EGFP) and HIV protein was used. The expression of the reporters can be easily visualized by fluorescence microscopy in living cells. siRNA-generating vectors targeted to several HIV-1 genomes were constructed and then co-transfected with respond reporter expression vectors into HEK293 cells. Cells transfected with pHIV-siRNA exhibited a significant inhibition of pEGFP-HIV expression compared with cells transfected with control vectors. By this way, it is successfully to select effective siRNA for silencing target HIV-1 genes. Then two or three siRNA transcripts targeted to different HIV genes were expressed by one plasmid, and a relative strong inhibition effect was observed.