Supplemental Material Materials and Methods

To characterize β-galactosidase-deficient Mariner transposon mutants, their total genomic DNA was isolated from overnight cultures with a GenElute Bacterial Genomic DNA Extraction Kit (Sigma-Aldrich, St. Louis, MO, USA) according to the manufacturer’s instructions. Genomic DNA (5 µg) was digested with RsaI (New England Biolabs, Ipswich, MA, USA) for four hours at 37C run on a 0.9% agarose gel in TAE buffer, and the DNA fragments of 1-4 Kb were excised from the gel and purified with the Illustra GFX PCR DNA/Gel Band Purification kit (GE Lifesciences, Buckinghamshire, UK). The purified DNA fragments were incubated with T4 DNA ligase (New England Biolabs) under the conditions that favor intra-molecular ligation at 14C overnight and resulting DNA circles were subjected to inverse-PCR (iPCR) using primers CJK94 5’- GGCTTGAACGAATTGTTAGGTGGC-3’ and CJK95 5’- CGGCCGCGTAATACGACTCACTA-3’ with the following cycle conditions (94C, 5 min; 35 cycles of 94C, 1 min; 53C, 1 min; 72C, 2 min; final extension at 72C, 10 min). iPCR products were cloned into pCR2.1-TOPO (Invitrogen, Carlsbad, CA, USA), transformed into chemically competent E. coli DH5α and sequenced with M13F/R primers at the University of Florida Biotechnology Core Facility.

Because plasmids containing S. marcescens PDL100 operon did not complement the phenotype of CK2A4, E. coli mutants in malE, malF and malG were constructed. Individual E. coli BW25113 mutants were obtained through the Keio Collection . As E. coli strain BW25113 is lac^- , each of the mutants was transduced into strain W3110 (lac⁺) using phage P1 . The resulting frt-kan-frt marker was excised using pCP20 and the plasmid was cured by incubation at 37°C . These mutants were used for the complementation experiments using pCJK17 and pCJK18 plasmids and for the phenotypic assays. Activities of β-galactosidase, N-acetyl-β-D-glycopyranosidase, and α-D-glucopyranosidase were tested in the wild type and mutant strains of both S. marcescens and E. coli .

In order to begin to determine the type of inhibitory compounds produced by Exigobacterium sp. 33G8 in co-culture, cell-free supernatant from the 33G8-PDL100 co-incubation experiment was extracted by flash ion exchange chromatography followed by C18 resin chromatography. At the time of extraction the OD₆₀₀ of the 33G8 culture was ~1.5. 500 ml of the culture were pelleted at 14,000 rpm for 10 minutes using an Avanti J-26 XP centrifuge (Beckman Coulter, Atlanta, GA, USA), the supernatant was transferred to a new centrifuge tube and centrifuged at 14,000 rpm for 10 minutes to remove all of the cells. The cell-free supernatant was applied to an ion exchange resin column (DOWEX-50W, Sigma-Aldrich, St. Louis, MO, USA). The flow-through was then subject to reverse phase Silica-C₁₈ flash chromatography (Alltech Associates, Inc., Deerfield, IL, USA) in sequence. For both chromatographies, resin bed volume was 10 ml. DOWEX-50W ion exchange resin was charged with 10 bed volumes of HPLC water prior to the application of the sample. The reverse phase Si-C₁₈ resin was first washed in reagent ethanol and then equilibrated in 10 bed volumes of HPLC water.