Ambion's new pSilencer adeno 1.0-CMV System combines the advantages of adenoviral vectors with an efficient siRNA expression system (see A Bit About Adenoviral Biology, at right). With pSilencer adeno, you can easily deliver an siRNA expression construct into a variety of mammalian cells and organisms. The modified CMV promoter in the vector efficiently expresses hairpin siRNAs, which elicit gene silencing through the RNAi pathway. Figures 1 and 2 show silencing of GAPDH and GFP in HeLa cells after infection with pSilencer adeno 1.0-CMV engineered to express the corresponding siRNAs. In both examples, protein levels were reduced ~90% compared to noninfected controls. The vector and promoter featured in the pSilencer adeno 1.0-CMV System were developed by Beverly Davidson and colleagues at the University of Iowa. This vector has been used by that lab to silence a number of genes in HeLa cells and adult mice (1).
Figure 1. Silencing of GAPDH Induced by pSilencer adeno. HeLa cells were infected at an MOI of 80 with adenovirus derived from the pSilencer adeno 1.0-CMV System. The virus was designed to express a GAPDH siRNA or a scrambled negative control siRNA. The media was changed 4 hours after infection and cells were analyzed after 72 hours. Immunofluorescence was performed using mouse anti-GAPDH antibody. Green: GAPDH protein. Blue: DAPI stained nuclei.
Figure 2. Silencing