ArticleBMC biotechnology2015
A simple and efficient seamless DNA cloning method using SLiCE from Escherichia coli laboratory strains and its application to SLiP site-directed mutagenesis.
Article in BMC biotechnology, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 122 papers.
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122 citing papers in PubMed, 223 citations in OpenAlex.
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62 more citing papers are in PubMed but not listed here.
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1 author at 1 institution in 1 country.
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Abstract
backgroundSeamless ligation cloning extract (SLiCE) is a simple and efficient method for DNA assembly that uses cell extracts from the Escherichia coli PPY strain, which expresses the components of the λ prophage Red/ET recombination system. This method facilitates restriction endonuclease cleavage site-free DNA cloning by performing recombination between short stretches of homologous DNA (≥ 15 base pairs).
resultsTo extend the versatility of this system, I examined whether, in addition to bacterial extracts from the PPY strain, other E. coli laboratory strains were suitable for the SLiCE protocol. Indeed, carefully prepared cell extracts from several strains exhibited sufficient cloning activity for seamless gene incorporation into vectors with short homology lengths (approximately 15-20 bp). Furthermore, SLiCE was applied to the polymerase chain reaction (PCR)-based site-directed mutagenesis method, in a process termed "SLiCE-mediated PCR-based site-directed mutagenesis (SLiP site-directed mutagenesis)". SLiP site-directed mutagenesis simplifies the steps of PCR-based site-directed mutagenesis, as it exploits the capability of the SLiCE method to insert multiple fragments.
conclusionsSLiCE can be performed in the laboratory with no requirement for a special E. coli strain, and the technique is easily established. This method increases the cloning efficiency, shortens the time for DNA manipulation, and greatly reduces the cost of seamless DNA cloning.
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