Abstract: To explore the effect of RNA interference on the suppression of goat prion protein gene (PRNP), small interfering RNA (siRNA) expressing vectors pSilencer4.1-siRNA1 and a negative control pSilencer4.1-siRNA2 and the eukaryotic expressing vector pSG5-prion expressing parts of goat prion protein gene were designed and constructed. The recombinants were introduced into Human Embryonic Kidney (HEK) 293T cells mediated by GeneTran. RT-PCR was carried out to detect the expression of prion protein gene. The results showed that RNAi effects induced by pSilencer4.1-siRNA1 could successfully deplete the mRNA expression of goat prion protein gene in 293T cells 48 h after transfection with both vectors compared to the negative control. The findings demonstrated that pSilencer4.1-siRNA1 was constructed successfully to transfer antagonistic substance against goat prion protein gene.