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| The EMBO Journal
(2001) 20, 3251–3261, doi:10.1093/emboj/20.12.3251 |
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| Product analysis illuminates the final steps of IES deletion in Tetrahymena thermophila |
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| Sergei V. Saveliev and Michael M. Cox |
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Department of Biochemistry, University of Wisconsin-Madison, 433 Babcock Drive, Madison, WI 53706-1544, USA To whom correspondence should be addressed Michael M. Cox, cox@biochem.wisc.edu Received 15 January 2001; Revised 20 April 2001; Accepted 20 April 2001. |
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| Abstract |
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| DNA sequences (IES elements) eliminated from the developing macronucleus in the ciliate Tetrahymena thermophila are released as linear fragments, which have now been detected and isolated. A PCR-mediated examination of fragment end structures reveals three types of strand scission events, reflecting three steps in the deletion process. New evidence is provided for two steps proposed previously: an initiating double-stranded cleavage, and strand transfer to create a branched deletion intermediate. The fragment ends provide evidence for a previously uncharacterized third step: the branched DNA strand is cleaved at one of several defined sites located within 15–16 nucleotides of the IES boundary, liberating the deleted DNA in a linear form. |
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| Keywords: ciliated protozoa, developmentally programmed genome rearrangements, DNA cleavage, DNA recombination, IES element |
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