An efficient method for generating human somatic cell gene knockouts.
Extract: The Human Genome Project has produced a map detailing a vast genetic frontier that will continue to provide useful insights for the treatment of human diseases. The large number of uncharacterized genes reflects the degree of our progress and the wealth of opportunity. Functional genomics will broadly impact our understanding of disease and illuminate the path to better therapeutics. One of the most definitive ways to determine gene function is to specifically inactivate a gene through knockout approaches, thereby permitting comparisons between genetically matched (i.e., isogenic) knockout and wild-type controls. Gene knockout technologies have been performed in a variety of model organisms, including bacteria, yeast, chickens, and rodents. Though these studies might be useful for inferring human gene function, it is clear that homologues are not always functionally identical. One of the best ways to study gene function in human cells is to generate a human somatic cell gene knockout. However, this approach has historically been inefficient, resulting in the widespread use of "knockdown" approaches employing antisense or RNA interference (siRNA, short interfering RNA) technologies. These approaches reduce, rather than eliminate, the expression of a particular gene and often have non-specific effects that complicate the analysis of gene function.
Kohli
Manu
M
Sidney Kimmel Comprehensive Cancer Center and The Cellular and Molecular Medicine Program, The Johns Hopkins University Medical Institutions, Baltimore, MD 21231, USA.
Rago
Carlo
C