Identification of effective siRNA sequence for RelB silencing in murine dendritic cells with siRNA cassette

Abstract

Abstract: Objective To construct small interfering RNA (siRNA) expression cassette targeting murine RelB gene and identify the most effective siRNA sequence against RelB gene in murine bone marrow-derived dendritic cells (DCs). Methods Three expression cassettes namely R1/siRNA, R2/siRNA and R3/siRNA targeting the sites 1027, 302 and 1121 of RelB gene, respectively, were constructed by PCR approach and transfected into cultured murine myeloid DCs by catione liposome AdvantGene. After incubation for 24 hours in a incubator containing 5% CO2 at 37 ℃, the DCs were stimulated by lipopolysaccharide (LPS), and RelB gene expression in DCs were then detected by RT-PCR and immunofluorescence. Results RT-PCR and immunofluorescence assay showed that the expression of RelB gene in DCs transfected with R2/siRNA could not be upregulated by LPS stimulation, but transfection with R1/siRNA or R3/siRNA failed to produce such effect. Conclusion R2/siRNA is an effective sequence for RelB silencing, and can be a useful means to construct new tolerogenic DC, RNAi RelB DC, for clinical immunotolerance induction.

CLC Number:

ZHENG Lei, BAO Jie, WANG Qian, YANG Hong-ling,QIU Yu-rong. Laboratory Medicine Center, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. Identification of effective siRNA sequence for RelB silencing in murine dendritic cells with siRNA cassette[J]. Journal of Southern Medical University, 2006, 26(03): 301-304.

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