Lysogeny broth agar

Manufactured by Merck Group

Sourced in United States, Canada, Singapore

Lysogeny broth (LB) agar is a nutrient-rich growth medium commonly used in microbiology and molecular biology laboratories. It provides the necessary nutrients and solidifying agent for the cultivation of a wide range of bacterial species. LB agar supports the growth of various microorganisms, enabling researchers to isolate and study them in a controlled environment.

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7 protocols using lysogeny broth agar

Drosophila Culture and Bacterial Strains

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D. melanogaster Canton-S flies were grown at 25° on a yeast glucose diet containing 100 g/ liter brewer’s yeast (inactive) (MP Biomedicals), 100 g/ liter glucose (Sigma), 12 g/ liter agar (Apex), and preservatives (0.04% phosphoric acid and 0.42% propionic acid (Sigma)) on a 12-h-light/12-h-dark cycle.
Bacterial strains used in the study are included in Table 1. Media used included lysogeny broth (LB)/agar (Sigma), modified MRS (mMRS) broth/agar and potato dextrose broth/ agar (Sigma). The plasmid bearing E. coli strain was cultured with 50 μg/ml kanamycin and Acetobacter gene knock-out mutants were cultured with 30 μg/ml chloramphenicol and 50 μg/ml kanamycin. All bacterial strains were cultured at 30°. A. fabarum DsW_054 was cultured in potato dextrose broth prior to matings.

White K.M., Matthews M.K., Hughes R.C., Sommer A.J., Griffitts J.S., Newell P.D, & Chaston J.M. (2018). A Metagenome-Wide Association Study and Arrayed Mutant Library Confirm Acetobacter Lipopolysaccharide Genes Are Necessary for Association with Drosophila melanogaster. G3: Genes|Genomes|Genetics, 8(4), 1119-1127.

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Bacterial Burden Evaluation Post-SCI

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At 7 days post-SCI, the bacterial burdens of bladder and urine were evaluated by plating serial dilutions of organ homogenates and urine onto Lysogeny broth (LB) agar (Sigma- Aldrich), a nutritionally rich medium and counting the colony-forming units to evaluate bacteria proliferation.

Campolo M., Siracusa R., Cordaro M., Filippone A., Gugliandolo E., Peritore A.F., Impellizzeri D., Crupi R., Paterniti I, & Cuzzocrea S. (2019). The association of adelmidrol with sodium hyaluronate displays beneficial properties against bladder changes following spinal cord injury in mice. PLoS ONE, 14(1), e0208730.

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Culturing and Characterizing Shiga Toxin-Producing E. coli Strains

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EC20020170 O113:H21, EC19990166 O145:H25, EC20010076 O91:H21, EC19970119 O26:H11, EC19990161 O121:H19, EC19960949 O128:NM, EC19970327 O103:H2, EC19940040 O45:H2, EC20030053 O111:NM, and EC2011007 O157:H7 were kindly provided by Dr. Roger Johnson of the Public Health Agency of Canada (Guelph, ON, USA). All strains were streaked onto Lysogeny broth (LB) agar (Sigma-Aldrich, Oakville, ON, USA) and incubated at 37 °C for 18 h. An isolated colony was then inoculated into 10 mL of Minimal Salt (M9) broth (Sigma-Aldrich) supplemented with 0.4% glucose, 0.02% MgSO₄, and 0.001% CaCl₂ (w/v) and grown at 37 °C, on a rocker platform at 180 rpm for 18 h.

Ma Z., Bumunang E.W., Stanford K., Bie X., Niu Y.D, & McAllister T.A. (2019). Biofilm Formation by Shiga Toxin-Producing Escherichia coli on Stainless Steel Coupons as Affected by Temperature and Incubation Time. Microorganisms, 7(4), 95.

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Multidrug-Resistant E. coli Biofilm Formation

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E. coli O154:H10 is a multidrug-resistant (streptomycin, tetracycline, ampicillin, chloramphenicol, trimethoprim-sulfamethoxazole, nalidixic acid and norfloxacin) strain isolated from South African cattle feces [7 (link)] and EC20020170. O113:H21 is a Canadian STEC, of human origin, provided by Dr. Roger Johnson of the Public Health Agency of Canada (Guelph, ON). These two strains were selected based on their strong biofilm-forming abilities on polystyrene [7 (link),73 (link)] and stainless-steel [8 (link),74 (link)]. The strains were streaked from glycerol stocks stored at −80 °C onto Lysogeny broth (LB) agar (Sigma-Aldrich, Oakville, Ontario, Canada) and incubated aerobically at 37 °C for 18 h.

Bumunang E.W., Ateba C.N., Stanford K., Niu Y.D., Wang Y, & McAllister T.A. (2020). Activity of Bacteriophage and Complex Tannins against Biofilm-Forming Shiga Toxin-Producing Escherichia coli from Canada and South Africa. Antibiotics, 9(5), 257.

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Bacterial Predation Assays with Bdellovibrio

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Lysogeny broth (LB) agar, agarose, glycerol, calcium chloride, magnesium chloride, polystyrene (PS), polycaprolactone (PCL), polylactic acid (PLA), and paraformaldehyde were obtained from Sigma‐Aldrich (Singapore). Mini hyaluronic acid (miniHA) powder was purchased from Bloomage Freda Biopharm Co., Ltd. (China). Phosphate‐buffered saline (PBS) and 4‐(2‐hydroxyethyl)‐1‐piperazineethanesulfonic acid (HEPES) were purchased from GE Hyclone (Singapore). FM 4–64FX dye was purchased from Thermo Fisher Scientific (Singapore). Nutrient broth and LB were purchased from BD Diagnostics (USA). B. bacteriovorus (ATCC 15143), E. coli (ATCC 25922), E. coli (ATCC 10536, only for B. bacteriovorus culturing), P. aeruginosa (PAO1‐GFP), A. baumannii (ATCC 19606), and K. pneumoniae (ATCC BAA‐2784) were purchased from ATCC (USA).

Cui M., Zheng M., Wiraja C., Chew S.W., Mishra A., Mayandi V., Lakshminarayanan R, & Xu C. (2021). Ocular Delivery of Predatory Bacteria with Cryomicroneedles Against Eye Infection. Advanced Science, 8(21), 2102327.

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Bacterial Growth Characterization

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Bacteria were plated out on Lysogeny Broth (LB) agar (Sigma) to single colonies prior to overnight growth in 5 ml liquid LB. 1:100 dilution of overnight culture was inoculated into 5 ml LB and growth assessed by optical density (OD₆₀₀). Bacterial growth was assessed in LB and M9 minimal salts media supplemented with 50 mM succinate or glucose where indicated.

Gill S.K., Hui K., Farne H., Garnett J.P., Baines D.L., Moore L.S., Holmes A.H., Filloux A, & Tregoning J.S. (2016). Increased airway glucose increases airway bacterial load in hyperglycaemia. Scientific Reports, 6, 27636.

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Preparation and Characterization of MWCNT

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All chemicals used in this study were of analytical grade and used as received.
Lysogeny broth (LB) agar, LB broth, and 50% Glutaraldehyde were purchased from Sigma. MWCNTs (724769, d: 6-9 nm with 5.5 nm mode, L: 5000 nm) were purchased from Aldrich. 10X phosphate buffered saline (PBS) stock solution was prepared by dissolving 1.3 M NaCl, 70 mM Na2HPO4.2H2O, and 30 mM NaH2PO4.2H2O and adjusting the pH to 7.0; 1x PBS solution was subsequently prepared by diluting the 10X PBS stock with Milli-Q ultrapure water. All aqueous media used were sterilized by autoclave at 121 o C for 15 min.

Hartono M.R., Kushmaro A., Chen X, & Marks R.S. (2018). Probing the toxicity mechanism of multiwalled carbon nanotubes on bacteria. Environmental science and pollution research international, 25(5).

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