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Nucleic Acids Research, 2002, Vol. 30, No. 7 e29
© 2002 Oxford University Press

An in vivo selection system for homing endonuclease activity

Mathias Gruen, Kathy Chang, Irina Serbanescu and David R. Liu*

Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA

Homing endonucleases are enzymes that catalyze the highly sequence-specific cleavage of DNA. We have developed an in vivo selection in Escherichia coli that links cell survival with homing endonuclease-mediated DNA cleavage activity and sequence specificity. Using this selection, wild-type and mutant variants of three homing endonucleases were characterized without requiring protein purification and in vitro analysis. This selection system may facilitate the study of sequence-specific DNA cleaving enzymes, and selections based on this work may enable the evolution of homing endonucleases with novel activities or specificities.

* To whom correspondence should be addressed. Tel: +1 617 496 1067; Fax: +1 617 496 5688; Email: drliu{at}fas.harvard.edu


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