Specific inhibition of OPGL expression by RNA interference

    DAI Xianª²Wen, WANG Quanª²Ping, YANG Min, LI Liª²Wen, HAN Liª²Hua, HU Yunª²Yu

    Institute of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi¡¯an 710033, China

    ¡¾Abstract¡¿ AIM: To construct the plasmid vector that can produce hairpin siRNA specific to rat osteoprotegerin ligand (OPGL) and to assess the inhibitory effects of this vector on OPGL mRNA. METHODS: Hairpin siRNA templates were designed based on rat OPGL gene sequence and mRNA structure and was cloned into mU6pro vector, which could produce hairpin siRNA to induce RNA interference in mammalian cells. The bone marrow stromal cells were transfected with the mU6proª²dsOPGL. After 48 hours, total RNA of cells was separated as samples for RTª²PCR with primers specific to OPGL and ¦Âª²actin. The difference of OPGL mRNA expression was assessed in view of the light density ratio of the RTª²PCR products of OPGL mRNA by gel analysis system. RESULTS: The plasmid vector with U6 promoter producing hairpin siRNAs to induce RNA interference was constructed successfully and the level of OPGL mRNA expression decreased with the increase of mU6proª²dsOPGL in bone marrow stromal cells. CONCLUSION: RNAª²interferenceª²induced inhibition of OPGL mRNA can significantly decrease the OPGL level, and the vector with U6 promoter producing siRNAs is responsible for the inhibition. ......