RT Journal Article SR Electronic(1) A1 Gleave, Andrew P. A1 Mountain, Andrew A1 Thomas, Christopher M.YR 1990 T1 Use of a novel cassette to label phenotypically a cryptic plasmid of Bacillus subtilis and map loci involved in its stable maintenance JF Microbiology, VO 136 IS 5 SP 905 OP 912 DO https://doi.org/10.1099/00221287-136-5-905 PB Microbiology Society, SN 1465-2080, AB In order to facilitate studies on the maintenance of cryptic plasmids from Gram-positive bacteria we have constructed a novel cassette cAPG1000 (5·0kb) which carries both a selectable marker (chloramphenicol resistance from Staphylococcus aureus plasmid pC194) and a screenable marker (the xylE gene from the TOL plasmid of Pseudomonas putida expressed from a cloned promoter of Bacillus phage SPO2) and which is flanked by terminators to prevent transcription from the cassette activating or inhibiting loci adjacent to the site of insertion. To demonstrate the usefulness of this cassette we have mapped loci required for stable maintenance of an 8·6 kb cryptic plasmid endogenous to Bacillus subtilis (pPOD2000) from the properties of cAPG1000 insertion and insertion/deletion derivatives. We have identified the replication region as well as separate regions required for segregational and structural stability. The segregational mechanism is very efficient since it allows no detectable loss despite the fact that bacteria carrying the plasmid have a greatly increased mean generation time., UL https://www.microbiologyresearch.org/content/journal/micro/10.1099/00221287-136-5-905