Figure 5.

pIJ303 Δ spdA2 and pIJ303-sps do not accumulate single-stranded plasmid DNA. Approximately 10⁶ protoplasts of S. lividans TK54 with respective plasmids were lysed in a 1% TA agarose gel containing 0.25% SDS (A). Black arrows indicate chromosomal DNA, while the gray arrow marks ccc-DNA. The gel was either blotted in native state (B) to detect accumulation of single-stranded plasmid DNA, or denatured before blotting in order to detect also double-stranded plasmid DNA (C). As a probe a 600 bp PCR-fragment corresponding to the dso of pIJ101 was used. Plasmid pIJ702, which lacks the sso, was used as a positive control for the accumulation of single-stranded plasmid DNA (white arrow). 1, pIJ303; 2, pIJ303-sps; 3, pIJ303Δ spdA2; 4, pIJ303spdA2c; 5, pIJ702; 6, plasmid free TK54.