Abstract
Mobile group II introns can be retargeted to insert into virtually any desired DNA target. Here we show that retargeted group II introns can be used for highly specific chromosomal gene disruption in Escherichia coli and other bacteria at frequencies of 0.1–22%. Furthermore, the introns can be used to introduce targeted chromosomal breaks, which can be repaired by transformation with a homologous DNA fragment, enabling the introduction of point mutations. Because of their wide host range, mobile group II introns should be useful for genetic engineering and functional genomics in a wide variety of bacteria.
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Acknowledgements
This work was supported by NIH grant GM37949. We thank Stephanie Reeves, Shelley Payne and Rasika Harshey for Shigella and Salmonella strains and Marlene Belfort and Keith Derbyshire (Albany) for comments on the manuscript.
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Karberg, M., Guo, H., Zhong, J. et al. Group II introns as controllable gene targeting vectors for genetic manipulation of bacteria. Nat Biotechnol 19, 1162–1167 (2001). https://doi.org/10.1038/nbt1201-1162
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DOI: https://doi.org/10.1038/nbt1201-1162
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