Use of bacteriophage lambda recombination functions to promote gene replacement in Escherichia coli.
TLDR
The deltarecBCD::Plac-red kan replacement allele can be P1 transduced to other E. coli strains, making the hyper-Rec phenotype easily transferable.Abstract:
Replacement of Escherichia coli's RecBCD function with phage lambda's Red function generates a strain whose chromosome recombines with short linear DNA fragments at a greatly elevated rate. The rate is at least 70-fold higher than that exhibited by a recBC sbcBC or recD strain. The value of the system is highlighted by gene replacement with a PCR-generated DNA fragment. The deltarecBCD::Plac-red kan replacement allele can be P1 transduced to other E. coli strains, making the hyper-Rec phenotype easily transferable.read more
Citations
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One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products
TL;DR: A simple and highly efficient method to disrupt chromosomal genes in Escherichia coli in which PCR primers provide the homology to the targeted gene(s), which should be widely useful, especially in genome analysis of E. coli and other bacteria.
Journal ArticleDOI
An efficient recombination system for chromosome engineering in Escherichia coli
TL;DR: A recombination system has been developed for efficient chromosome engineering in Escherichia coli by using electroporated linear DNA using a defective lambda prophage, which will be especially useful for the engineering of large bacterial plasmids such as those from bacterial artificial chromosome libraries.
Journal ArticleDOI
PCR-targeted Streptomyces gene replacement identifies a protein domain needed for biosynthesis of the sesquiterpene soil odor geosmin
TL;DR: An efficient procedure for creating precise gene replacements in the cosmid clones by using PCR targeting and λ-Red-mediated recombination is described, which is used successfully by >20 researchers to mutate around 100 Streptomyces genes.
Journal ArticleDOI
A new logic for DNA engineering using recombination in Escherichia coli
TL;DR: A straightforward way to engineer DNA in E. coli using homologous recombination is described in this article, which uses RecE and RecT and is transferable between different E coli strains.
Journal ArticleDOI
The biofilm matrix – an immobilized but dynamic microbial environment
TL;DR: Although exopolysaccharides provide the matrix framework, a wide range of enzyme activities can be found within the biofilm, some of which will greatly affect structural integrity and stability.
References
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A novel suicide vector and its use in construction of insertion mutations: osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR.
TL;DR: Certain environmental signals (i.e., osmolarity and the presence of amino acids) are tightly coupled to the expression of toxR-regulated proteins and therefore may be signals that are directly sensed by the ToxR protein.
Journal ArticleDOI
New method for generating deletions and gene replacements in Escherichia coli.
TL;DR: A method for generating gene replacements and deletions in Escherichia coli using a temperature-sensitive pSC101 replicon to facilitate the gene replacement and can be used to generate deletions of essential genes.
Journal ArticleDOI
Selection for loss of tetracycline resistance by Escherichia coli.
Stanley Maloy,W D Nunn +1 more
TL;DR: An improved medium for the direct, positive selection of tetracycline-sensitive clones from a population of t Petracy Cline-resistant strains of Escherichia coli is described.