Group II introns as controllable gene targeting vectors for genetic manipulation of bacteria
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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The intron-based approach utilizes the specificity of mobile group II intron Ll.ltrB from Lactoccocus lactis to propagate into the specified site via a RNA-mediated, retro-homing mechanism.
“The approach utilizes the specificity of mobile group II intron Ll.ltrB from Lactoccocus lactis to propagate into the specified site via a RNA-mediated, retro-homing mechanism (Karberg et al., 2001).”
From Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks (2016) · cites this paper as Karberg et al., 2001
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- Authors
- Karberg, M., Guo, H., Zhong, J., Coon, R., Perutka, J., & Lambowitz, A. M.
- Affiliations
- The University of Texas at Austin (US)
- Venue
- Nature Biotechnology
- Published
- 2001-12
- Volume
- 19
- Issue
- 12
- Pages
- 1162-1167
- ISSN
- 0733-222X (ISSN-L)
- 1087-0156
- 1546-1696
- OpenAlex
- W2060576378
- Cited by
- 222
- References
- 29
- Citations by year
- 2026: 5 · 2025: 4 · 2024: 7 · 2023: 2 · 2022: 4 · 2021: 10 · 2020: 3 · 2019: 12 · 2018: 10 · 2017: 3 · 2016: 10 · 2015: 11 · 2014: 21 · 2013: 16 · 2012: 11
- FWCI
- 2.44
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Nature Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institute of General Medical Sciences (GM37949, R01GM037949)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
- Keywords
- group II introns, gene targeting, Escherichia coli, homologous recombination, point mutations, functional genomics, mobile genetic elements
- MeSH terms
- Base Sequence, Chromosomes, Escherichia coli, Genes, Bacterial, Genetic Engineering, Genetic Vectors, Introns, Molecular Sequence Data, Plasmids, Recombination, Genetic, Blotting, Southern, Gene Library, Polymerase Chain Reaction, Point Mutation, Gene Transfer Techniques, Codon, Terminator
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