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Nucleic Acids Research, 2003, Vol. 31, No. 14 3993-4000
© 2003 Oxford University Press

The frequency of gene targeting in Trypanosoma brucei is independent of target site copy number

Bill Wickstead, Klaus Ersfeld and Keith Gull*

School of Biological Sciences, University of Manchester, 2.205 Stopford Building, Oxford Road, Manchester M13 9PT, UK

*To whom correspondence should be addressed at present address: Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK. Tel: +44 1865 285 455; Fax: +44 1865 285 691; Email: keith.gull{at}pathology.oxford.ac.uk
Present address:
Bill Wickstead, Sir William Dunn School of Pathology, University of Oxford, South Parks Road, Oxford OX1 3RE, UK

We have investigated the effect of target copy number on the efficiency of stable transformation of the protozoan parasite Trypanosoma brucei. Using a single strain of the organism, we targeted integrative vectors to several different genomic sequences, occurring at copy numbers ranging from 1 to ~30 000 per diploid genome, and undertook a systematic assessment of both transformation and integration efficiencies. Even over this vast copy number range, frequency of gene targeting was the same for all sites. An independence of targeting frequency and target copy number is characteristic of mammalian homologous recombination and is unlike the situation in budding yeast. It is also not seen in the related parasite Leishmania, a distinction that may be the consequence of the different usage of recombination within the mechanisms of pathogenicity in the two species.


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