Journal: The ISME Journal

Article Title: A new biofilm-associated colicin with increased efficiency against biofilm bacteria

doi: 10.1038/ismej.2013.238

Figure Lengend Snippet: ROAR029 bacteriotoxic activity is due to plasmid-encoded colicin regulated by PcnB. (a) Addition of an 8-μl drop of ROAR029 wild type, ROAR029ΔpcnB or ROAR029 bearing a mutagenized plasmid unable to synthesize the bacteriotoxic molecule (ROAR029 pColR−) biofilm extract to a lawn of MG1655. (b) DNA gel of ROAR029 pColR plasmid linearized by the EcoRV enzyme. (c) ROAR029 pColR plasmid map. Arrows indicate transcription polarities of colicin genes. Unique-site restriction enzymes are indicated. (d) Dendrogram of sequences alignment of the 200 last amino acids from colicin-encoding genes of the C-terminal region. Phylogenetic analysis was carried out using the UPGMA method. Numbers represent bootstrap values >75.

Article Snippet: Plasmid map pColR plasmid map was constructed using the SnapGene viewer from GSL Biotech LLC (Chicago, IL, USA).

Techniques: Activity Assay, Plasmid Preparation

Journal: The ISME Journal

Article Title: A new biofilm-associated colicin with increased efficiency against biofilm bacteria

doi: 10.1038/ismej.2013.238

Figure Lengend Snippet: ROAR029 biofilm-specific production of colicin R is due to an SOS regulatory effect. (a) Sequence analysis revealed two overlapping SOS boxes upstream of the ribosome binding sequence (rbs) and start codon ATG of ROAR029. Insertion site of transposon in pColR+ plasmid is indicated. Transcriptional read-through from kanamycin gene and disruption of LexA box1 binding site may explain constitutive expression of colicin R encoding gene. (b) Effect of LexA induction upon ROAR029 bacteriotoxic activity was assessed using ROAR029 with LexA under the control of an anhydrotetracycline-inducible (aTc) promoter. An 8-μl drop of ROAR029_KmRExTET_LexA treated with increasing concentrations of aTc ranging from 0 to 1000 ng ml−1 was added to a lawn of MG1655. (c) Inhibition halo of E. coli MG1655 when treated with an 8-μl drop of ROAR029 biofilm extract (ROAR029 Bf) with and without induction of the SOS system by 0.1 μg ml−1 of mitomycin C added after inoculation. (d) Growth inhibition of E. coli MG1655 by biofilm extracts produced by E. coli TG1 (black) and TG1 lexAind3 (gray) transformed by different ROAR029 plasmids; active kanamycin-resistant pColR_Km, defective colicin mutant (pColR-, dashed) and constitutively expressed colicin (pColR+, shaded).

Article Snippet: Plasmid map pColR plasmid map was constructed using the SnapGene viewer from GSL Biotech LLC (Chicago, IL, USA).

Techniques: Sequencing, Binding Assay, Plasmid Preparation, Disruption, Expressing, Activity Assay, Control, Inhibition, Produced, Transformation Assay, Mutagenesis