sentence: A 47 bp fragment of the comN ORF and the scar of the cre ‐ lox system were left behind encoding the following peptide : VEKHPADPTVRIAYIIRTVGSQEALEELLR.A minD deletion mutant ( marker ‐ free ) was constructed by growing strain Δ minD : : spec P ‐ mazF two consecutive times on NA plates containing 1 mM IPTG and selecting for spectinomycin ‐ sensitive clones.A P ‐ comN ‐ gfp fusion was made by cloning a PCR product encompassing P ‐ comN , which was generated from B. subtilis 168CA chromosomal DNA with primers oTK10 / oTK11 into the SphI , KpnI digested pSG1154mgfp [ a monomeric GFP variant made by Henrik Strahl based on pSG1154 of ( Lewis and Marston , 1999 ) ] , thereby removing P .