Fig. 2

General diagram representing the generic method for gene modification by allelic exchange in solventogenic Clostridia using a restrictionless ∆upp strain. The example present the use of the method for markerless gene deletion, but it can also be used for markerless gene editing or DNA insertion. The gene to delete in frame is y. The boxed regions of x and z genes represent approximatively the regions of homology incorporated into the suicide plasmid. a Selection for plasmid integration in 5′ or 3′ using the antibiotic resistance carried by the suicide vector. b Counter selection strategy with the 5-FU/upp system used for the selection of the double crossing-over and the excision of the plasmid