Stimulation of homologous recombination in plants by expression of the bacterial resolvase RuvC

Targeted gene disruption exploits homologous recombination (BR) as a powerf ul reverse genetic tool, for example in bacteria, yeast, and transgenic kno ckout mice, but it has not been applied to plants, owing to the low frequen cy of HR and the lack of recombinogenic mutants. To increase the frequency of HR in plants,,ve constructed transgenic tobacco lines carrying the Esche richia coil RuvC gene fused to a plant viral nuclear localization signal. W e show that RuvC, encoding an endonuclease that binds to and resolves recom bination intermediates (Holiday junctions) is properly transcribed in these lines and stimulates HR We observed a 12-fold stimulation of somatic cross over between genomic sequences, a Ii-fold stimulation of intrachromosomal r ecombination, and a 56-fold increase for the frequency of extrachromosomal recombination between plasmids cotransformed into young leaves via particle bombardment. This stimulating effect may be transferred to any plant speci es to obtain recombinogenic plants and thus constitutes an important step t oward gene targeting.