Abstract
Recently, gene targeting enables site-directed mutagenesis in the mammalian genome. It utilizes homologous recombination between DNA sequences of a targeting vector and the genome of mouse ES cells. Gene targeting conventionally achieves the disruption of the gene, or “gene knock-out”, inserting the drug resistant gene sequence not only as a selected marker, but also as an insertion sequence into the target gene by homologous recombination. Subsequent germ-line transmission of the ES cells through injection chimeras has contributed to the studies on the biological function of the disrupted gene in vivo.
Biological studies to create new drugs for human diseases could be furthermore elaborated if we were able to replace a target gene with human homologue. For this purpose, we have devised a new method that is able to replace the target gene with any sequences constructed in the replacement vector; including subtle mutations, deletions, insertions, human homologues, or totally non-homologous fragments. We conclude that this method, “universal gene replacement”, now allows us to alter any part of mammalian genome as desired DNA sequences of the target locus and it could be developed to a potent method for the generation of the humanized animal models.
