Sigma-Aldrich introduces gene knockout system
that is set to feature prominently in functional genomics and
systems biology. It incorporates technology that exploits the
retrohoming ability of group II introns, which "target" the exact
position of gene disruption.
TargeTron provides targeted, stable, permanent gene knockout. It is a simple to use technology that provides knockouts in 3 days and requires minimal screening of colonies to isolate mutants.
The method uses the retrohoming ability of group II introns and utilises a PCR step to "re-target" the TargeTron group II intron for specific insertion into the host genome.
A spokesman for Sigma-Aldrich told DrugResearcher.com that they had identified a segment of the drug discovery market that looked at prokaryotic organisms for potential drug targets.
"Targetron can be used in gene knockout studies and manipulation of metabolic pathways in prokaryotes," he added.
Licensed in Sept of 2004, Sigma has been developing the kit since. The technology allows permanent gene disruption to be performed in a non-random or targeted manner that does not require host recombination factors to mobilise.
An extensively validated TargeTron algorithm designs specific primer sets for use in PCR mutation of the TargeTron Intron. The mutated intron is essentially "re-programmed" to insert itself into the algorithm predicted site of any user-defined gene.
Ligation of the PCR product into a TargeTron vector is followed by transformation of the host cell. Expression results in a ribonucleoprotein complex that targets the intron to the insertion site within the host genome resulting in gene knockout.
A spokesman for Sigma Aldrich told DrugResearcher.com: "TargeTron offers a solution to gene knockout technologies. We saw large potential sales in the existing system as well as the possibility of expanding into other model organisms."
"We did see the technology as filling a "gap" in the market in terms of providing targeted, stable knockouts as well as the potential to perform functional genomics studies on difficult organisms such as clostridium perfringens."
The technology has provided greater than 90 per cent successfully targeted insertion rate in prokaryotes. The current TargeTron system has been used to create permanent knockouts across a broad range of bacterial strains such as Escherichia coli, Staphylococcus aureus, Lactococcus lactis, Clostridium perfringens, Shigella flexneri and Salmonella typhimurium. Future systems will be developed for both plant and mammalian-targeted gene disruption.
The current kit makes it ideal for academic pharmaceuticals, biotechnology companies and government-sponsored projects. The product has applications in prokaryotic functional genomics, systems biology and geneic engineering.
The kit has been designed for gene disruptions primarily in E. coli and other taxonomically related bacteria. Modification to the vector and promoters may be required to express the group II intron RNA-protein complex (RNP) in other bacterial strains.
"Group II intron-mediated gene disruption offers advantages over gene knockout techniques such as randomly integrated transposons and cumbersome recombination methods," commented Keith Jolliff, global marketing manager of molecular biology at Sigma-Aldrich.
Current technologies require more complicated and laborious practices requiring large constructs that are difficult to manipulate. Traditional methods also require the screening of thousands of colonies to isolate a knockout mutant.
Sigma-Aldrich's kit launches enter a sector that is currently experiencing a period of prosperity. Indeed functional genomics and systems biology is a very broad market that is experiencing large growth due to the introduction and widescale acceptance of siRNA technology.
According to Kalorama, RNAi/siRNA technology is currently the "hot property" in NA-based therapeutics, lthough it is still very much in its early stages of development with obstacles to its commercialisation yet to be overcome.
Meanwhile Frost and Sullivan's analysts estimate target validation to generate $146.4 million (€112.5 million) or just under 50 per cent of total RNAi market revenues, followed by research with $97.6 and therapeutics with $56.0 million by 2010.
TargeTron is available from Feb 15 and is available in two kits. TA0100-3ea is $300 and TA0100-10ea is $750.
