¹Division of Medicine, UCL Medical School, London, UK

Correspondence: Dr JS Owen, Division of Medicine (Upper 3rd Floor), UCL Medical School, Royal Free Campus, Rowland Hill Street, London NW3 2PF, UK. E-mail: j.owen@medsch.ucl.ac.uk

²Current address: Gene Targeting Group, Department of Haematology, Faculty of Medicine, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 ONN, UK.

Received 23 October 2008; Revised 27 February 2009; Accepted 28 February 2009; Published online 2 April 2009.

Abstract

Several independent groups have reported targeted genomic editing in mammalian cells mediated by synthetic oligonucleotides. Nevertheless, the validity of data has been disputed because of experimental artefacts, inconsistent findings and low reproducibility. Here, we describe experiments designed to meet stringent criteria and completely eliminate artefactual results. In particular, by targeting cells expressing mutated enhanced green fluorescence protein (EGFP), which allow editing measurements at the protein level, and analyzing corrected clones by Southern blotting, we rigorously excluded spontaneous reversion, contamination artefacts, false-positives, or overestimation. Our findings provide unequivocal authentication that oligonucleotide-mediated gene editing is a real, not artefactual, phenomenon—a vital starting point from which to develop the technology into practical applications.