Jb. Scheerer et Gm. Adair, HOMOLOGY DEPENDENCE OF TARGETED RECOMBINATION AT THE CHINESE-HAMSTER APRT LOCUS, Molecular and cellular biology, 14(10), 1994, pp. 6663-6673
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.