FIG 4.
Design of a plasmid that can be transferred to cells via conjugation or CNPX-mediated transduction. (A) Schematic of transduction and conjugation of pC221-derived plasmids into a host carrying a targeting CRISPR-Cas system. (B) Mobilizable, tetracycline-resistant pC221-pT181 hybrid plasmid used in this study. It was created by fusion of the mobilization genes (orfA and orfB) and origin of transfer (oriT) of pC221 with the tetracycline resistance (Tetr) gene, replication gene (repC), and recombination gene (pre) of pT181, as well as its origin of replication gene (ori). The site of target insertion is shown. (C) Schematic of the pTgt plasmid series showing the insertion of the nes or cn20 targets in the leading or lagging strand, according to replication from the ori site of pT181.