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Superior Gene Silencing Using Adenoviral Vectors


Mediating RNA interference (RNAi) with recombinant adenovirus expressing siRNAs has only recently been described in the literature (1). Recombinant adenoviruses have several advantages over other methods of nucleic acid delivery. Namely, these viruses can infect dividing and non-dividing cells, and can be used in animal model systems. They can infect many difficult-to-transfect primary and transformed cell lines via their receptor mediated delivery mechanism. The sequence of a chemically synthesized siRNA capable of gene specific silencing can also be functionally expressed from recombinant adenovirus, often at a lower cost. Once made, the virus can be expanded, making it possible to generate "stocks" for future experiments.


How Does pSilencer Adeno 1.0-CMV Work?
An oligonucleotide insert encoding the desired siRNA is cloned into the pre-linearized shuttle vector (provided), which includes a modified CMV promoter for efficient expression of the siRNA sequence. The shuttle vector, containing the desired sequence, is then linearized and introduced into HEK 293 packaging cells (not included) along with the linearized adenovirus backbone (Figure 1). The HEK 293 packaging cells contain the EIA and EIB genes which are not present in the introduced vectors and are required to form infectious particles. Inside the HEK 293 cells, the shuttle vector and the adenoviral backbone plasmid recombine to produce an adenoviral genome including the siRNA expression cassette. The siRNA expression cassette is packaged into infectiou
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