Turning off genes to elucidate their function.
Deleting a relevant mammalian gene constitutively (in the germ line) or conditionally (in certain tissues or upon intervention) is a way to investigate its malfunction or its potential as a drug target in a variety of disease areas – in neurodegeneration, cancer, CVD, and many other fields.
Since the advent of easily accessible techniques for conditional knockouts in mice, constitutive KO’s were more rarely generated. Currently, we generate constitutive knockouts predominantly for species that are less easily accessible – rats and rabbits. Here, a constitutive knock out is possible via nuclease based engineering (CRISPRs, TALENs, etc.).
Here’s when we target the gene in a manner that leaves it intact and fully functional, but allows for conditional inactivation upon combining with publicly available mouse strains that are pre-validated for this purpose. Conditional targeting also allows to irreversibly (“constitutively”) disrupt the gene in the germ line, upon appropriate breeding.
At PolyGene we do conditional knockouts in mice and rats via the CRISPR/Cas9 technology. Please in inquire for more details.
We flank a region of interest with loxP sites; we do so by targeting that region with a recombination vector containing the loxP sites. Typically, a few relevant exons and up to 2 kbp are flanked. Due to specialized base vectors, construction is almost as easy, and as rapid, as building a constitutive targeting vector.