HOMOLOGY DEPENDENCE OF TARGETED RECOMBINATION AT THE CHINESE-HAMSTER APRT LOCUS

Using simple linear fragments of the Chinese hamster adenine phosphori bosyltransferase (APRT) gene as targeting vectors, we have investigate d the homology dependence of targeted recombination at the endogenous APRT locus in Chinese hamster ovary (CHO) cells. We have examined the effects of varying either the overall length of targeting sequence hom ology or the length of 5' or 3' banking homology on both the frequency of targeted homologous recombination and the types of recombination e vents that are obtained. We find an exponential (logarithmic) relation ship between length of APRT targeting homology and the frequency of ta rgeted recombination at the CHO APRT locus, with the frequency of targ eted recombination dependent upon both the overall length of targeting homology and the length of homology flanking each side of the target gene deletion. Although most of the APRT(+) recombinants analyzed refl ect simple targeted replacement or conversion of the target gene delet ion, a significant fraction appear to have arisen by target gene-templ ated extension and correction of the targeting fragment sequences. APR T fragments with limited targeting homology flanking one side of the t arget gene deletion yield proportionately fewer target gene conversion events and proportionately more templated extension and vector correc tion events than do fragments with more substantial flanking homology.