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Allele-Coupled Exchange for chromosomal manipulation of Clostridioides difficile ribotype 027 and other ribotypes. Uses inducible expression of CD2517.1 toxin as a counter-selection marker
ColE1 ori, KanR, TetR, Tet promoter with a codon optimized pbTTA gene bearing an N-terminal hexahistidine tag followed by a SUMO tag and a TEV protease cleavage site.
SB-transposon with inducible SfiI cloning site for GOI (contains firefly luciferase) and constitutive expression of BFP, rtTA and hygromycin resistance gene