The effect of target site transcription on gene targeting in human cells in vitro

Bhaskar Thyagarajan, Brad L. Johnson, Colin R Campbell

Research output: Contribution to journalArticlepeer-review

39 Scopus citations

Abstract

We evaluated the effect of target site transcription on gene targeting In cultured human flbrosarcoma cells. A number of cell lines that harbored a plasmid recombination substrate within their chromosomal DNA were created. Gene targeting frequency was then measured at these different loci in the presence and absence of an agent that stimulated target site transcription. We observed that gene targeting was significantly enhanced by RNA transcription. The magnitude of transcription-stimulated gene targeting varied from 3-fold to >20-fold. No increase in gene targeting was observed, however, when transcription proceeded away from, rather than through, the recombination site. Transcription-stimulated gene targeting was also observed when single-stranded plasmid vectors complementary to either the coding or template strand were used as recombination substrates. Our results indicate that gene targeting, like other forms of DNA recombination, can be stimulated by target site transcription. The implications of our observations on current models of transcription-stimulated recombination are discussed.

Original languageEnglish (US)
Pages (from-to)2784-2790
Number of pages7
JournalNucleic acids research
Volume23
Issue number14
DOIs
StatePublished - Jul 25 1995

Bibliographical note

Funding Information:
This work was supported by a grant from the Minnesota Medical Foundation.

Copyright:
Copyright 2010 Elsevier B.V., All rights reserved.

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