Electrotransformation of unmethylated pDCW89 into JWCB018. (A) JWCB005 (ΔpyrFA) and JWCB018 (ΔpyrFA ΔcbeI) were transformed with un-methylated pDCW89 DNA and plated onto defined medium either with or without uracil. Controls with no DNA transformation are also presented. (B) Restriction analysis of pDCW89 plasmid DNA before and after transformation of C. bescii and back-transformation to E. coli. Lane 1, pDCW89 plasmid DNA isolated from E. coli DH5α, digested with either EcoRI (5.8 kb and 1.9 kb cleavage products), or with HindIII (4.4 kb and 3.3 kb cleavage products). Lanes 2, 3 and 4 plasmid DNA isolated from three biologically independent E. coli DH5α back-transformants using total DNA isolated from C. bescii transformants, digested with either EcoRI or HindIII. M: 1 KB DNA ladder (NEB).