Journal article

From cloning to mutant in 5 days: rapid allelic exchange in Staphylococcus aureus.

Ian R Monk, Timothy P Stinear

Access Microbiol | Microbiology Society | Published : 2021

Abstract

In the last 10 years, the barriers preventing the uptake of foreign DNA by clinical Staphylococcus aureus isolates have been identified and powerful mutagenesis techniques such as allelic exchange are now possible in most genotypes. However, these targeted approaches can still be cumbersome, and the construction of unmarked deletions/point mutations may take many weeks or months. Here, we introduce a streamlined allelic exchange protocol using IMxxB Escherichia coli and the plasmid pIMAY-Z. With this optimized approach, a site-specific mutation can be introduced into S. aureus in 5 days, from the start of cloning to isolation of genomic DNA for confirmatory whole-genome sequencing. This stre..

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