Nucleic Acids Research, 2002, Vol. 30, No. 22 4892-4901
© 2002 Oxford University Press
A chromosomal position effect on gene targeting in human cells
Gene Targeting Group, MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK
*To whom correspondence should be addressed. Tel: +44 20 8383 8276; Fax: +44 20 8383 8303; Email: andy.porter{at}csc.mrc.ac.uk
Present address: Rafael J. Yáñez, Molecular Immunology Unit, Institute of Child Health, University College London, 30 Guilford Street, London WC1N 1EH, UK
We describe gene targeting experiments involving a human cell line (RAN10) containing, in addition to its endogenous alleles, two ectopic alleles of the interferon-inducible gene 6-16. The frequency of gene targeting at one of the ectopic 6-16 alleles (H3.7) was 34-fold greater than the combined frequency of gene targeting involving endogenous 6-16 alleles in RAN10. Preference for H3.7 was maintained when the target loci in RAN10 were transcriptionally activated by interferon. Despite the 34-fold preference for H3.7, the absolute gene targeting efficiency in RAN10 was only 3-fold higher than in the parental HT1080 cell line. These data suggest that different alleles can compete with each other, and perhaps with non-homologous loci, in a step which is necessary, but not normally rate-limiting, for gene targeting. The efficiency of this step can therefore be more sensitive to chromosomal position effects than the rate-determining steps for gene targeting. The nature of the position effects involved remains unknown but does not correlate with transcription status, which in our system has a very modest influence on the frequency of gene targeting. In summary, our work unequivocally identifies a position effect on gene targeting in human cells.
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