Abstract
A two-plasmid mutagenesis system for Clostridioides difficile is described that improves ease of use and efficiency in creating site-directed mutations. pJB06 contains a xylose-inducible cas9 gene, while the second plasmid (pJB07) encodes the corresponding guide RNA (gRNA) and regions of homology for repair of the introduced double-stranded DNA (dsDNA) breaks, both of which are replaceable via restriction digest.
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