Bio-Rad keeps your genes silenced for longer
The web-based search tool, designed to save researchers time and effort is available at www.bio-rad.com/, and offers a way to review Bio-Rad's library of validated siRNA gene targets by searching for a target gene symbol, NCBI accession number, catalogue number, or browsing according to the area of interest.
The current research panels include areas such as angiogenesis, apoptosis, carcinogenesis, cell cycle/checkpoint, chromatin regulation, inflammation, kinases, metabolism and toxicology/ADME (absorption, distribution, metabolism, and excretion). The firm expects to add additional panels such as stem cell research, DNA damage/ repair, cardiovascular disease later this year.
According to Christina Whitman, product manager for Bio-Rad's RNAi / Gene Transfer Reagents in Bio-Rad's Gene Expression Division, the key advantages of using Bio-Rad's validated siLentMer siRNA products is the increased longevity of the gene silencing effect that can last up to nine days, compared with just three or four using traditional siRNAs.
The siLentMer reagents are based upon a discovery made by Dr John Rossi and Dr Mark Behlke of Integrated DNA Technologies in 2005 that showed that duplexes made of 27 nucleotides (27-mers) could be up to 100 times more potent than the commonly used 21-mers.
Prior to this discovery it had been believed that only 21-23 nucleotide double stranded siRNAs were believed to activate the RNA interference (RNAi) pathway in mammalian cells.
However, Rossi and Behlke showed that 27-mers could be designed so that they are precisely cleaved by the Dicer endonuclease into the active 21-mers which associate with an RNA-induced silencing complex (RISC) that targets homologous mRNA for degradation.
"The Dicer-substrate siRNAs can be up to 100 times more potent than the related 21-mer siRNA targeting the same sequence and can be active at concentrations as low as 100 pM for some gene targets which helps minimise the potential for off-target effects," said Whitman.
"Additionally, they can silence genes for twice as long as normal siRNAs. While 21-mers tend to knockdown a gene for three or four days, the Dicer substrate siRNAs can silence genes for up to 9 days, reducing the number of transfection steps needed when looking for effects of longer silencing, such as a phenotypic response."
The siLentMer siRNAs can be delivered into cells using a variety of methods, the most common of which use lipid-based delivery reagents. Other methods include electroporation and 'bioloistic' particle bombardment.
"Delivery is the most important step - it doesn't matter how good the siRNA sequence is, if you can't get it in the cell it's not going to work," said Whitman.
"This is why we've put a lot of effort into developing our delivery optimisation kits which contain fluorescently labelled nonsilencing 27-mer siRNA molecules that can be used to establish the optimal delivery conditions for any cell line."