May 30, 2000Proceedings of the National Academy of SciencesOpen Access

One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products

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Authors

KDKirill A. DatsenkoRutgers, The State University of New Jersey BWBarry L. WannerHarvard University

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Overview

Molecular analysis demonstrates efficient one-step chromosomal gene disruption in Escherichia coli using PCR products, highlighting a streamlined approach for bacterial genome editing.

Key Points

Chromosomal gene disruption was achieved across 13 target operons in Escherichia coli using PCR primers carrying short flanking homology, yielding antibiotic-resistant mutants.
Recombination using phage lambda Red recombinase facilitated targeted insertion of cassette DNA flanked by FRT sites, followed by FLP recombinase-mediated marker excision.
Supports high-throughput functional genomics across wild-type bacterial strains, eliminating complex cloning steps and enabling rapid chromosomal gene inactivation workflows.

Cite This Study

Datsenko et al. (2000) studied this question.

synapsesocial.com/papers/68f644b133dca03d982ea46c https://doi.org/10.1073/pnas.120163297

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May 30, 2000Proceedings of the National Academy of SciencesOpen Access

One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products

View Full Paper

Ask AI

Bookmark

Authors

KDKirill A. DatsenkoRutgers, The State University of New Jersey BWBarry L. WannerHarvard University

Discussion

Member takes

Overview

Molecular analysis demonstrates efficient one-step chromosomal gene disruption in Escherichia coli using PCR products, highlighting a streamlined approach for bacterial genome editing.

Key Points

Chromosomal gene disruption was achieved across 13 target operons in Escherichia coli using PCR primers carrying short flanking homology, yielding antibiotic-resistant mutants.
Recombination using phage lambda Red recombinase facilitated targeted insertion of cassette DNA flanked by FRT sites, followed by FLP recombinase-mediated marker excision.
Supports high-throughput functional genomics across wild-type bacterial strains, eliminating complex cloning steps and enabling rapid chromosomal gene inactivation workflows.

Cite This Study

Datsenko et al. (2000) studied this question.

synapsesocial.com/papers/68f644b133dca03d982ea46c https://doi.org/10.1073/pnas.120163297

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