Escherichia coli XL1 (Stratagene, USA) was used for maintenance of plasmid constructs and E. coli Rosetta BL21 pLysE (Merck Millipore, UK) was used for protein expression. E. coli strains were grown in Luria Bertani (LB) broth or agar at 37 °C, unless otherwise indicated, with shaking at 200 rpm for liquid cultures. C. jejuni M1 was used as a source of DNA for gene cloning and as the challenge strain in vaccination experiments as described [12] (link). C. jejuni 11168H was used to assess the cross-reactivity and subcellular localisation of SodB. C. jejuni strains were grown on modified charcoal-cephoperazone-deoxycholate agar (mCCDA) (Oxoid, UK) or in Mueller-Hinton Broth (MH; Oxoid), at 37 °C in a microaerophilic workstation (Don Whitley Scientific, UK) in a low oxygen atmosphere (5% O2, 5% CO2 and 90% N2). Liquid cultures of Campylobacter were grown with shaking at 400 rpm using a table top shaker (IKA, Germany) under low oxygen conditions as above. Antibiotics were used at the final concentrations of 100 μg/ml ampicillin and 34 μg/ml chloramphenicol where appropriate.
Mueller hinton broth mh
Manufactured by Thermo Fisher Scientific
Sourced in United Kingdom
Mueller-Hinton Broth (MH) is a laboratory culture medium used for the growth and isolation of various microorganisms. It is primarily used for antimicrobial susceptibility testing, such as the Kirby-Bauer disk diffusion method.
Automatically generated - may contain errors
Lab products found in correlation
Microaerophilic workstation, by Don Whitley Scientific (1 mentions)
Mccda, by Thermo Fisher Scientific (1 mentions)
Tryptic soy broth (tsb), by Thermo Fisher Scientific (1 mentions)
Sheep blood agar, by Thermo Fisher Scientific (1 mentions)
Ccda selective supplement, by Thermo Fisher Scientific (1 mentions)
Microbank vial, by Pro-Lab Diagnostics (1 mentions)
4 protocols using mueller hinton broth mh
1
Cultivation of E. coli and Campylobacter
2
Antibiotic Susceptibility of Environmental Bacteria
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The MIC of gentamycin and kanamycin against bacterial isolates representing every taxa identified by Sanger sequencing were measured based on a modification of the standard procedures given in CLSI (M100 2014). Five to ten different isolates from the three most abundant bacterial genera were analyzed. Twofold dilution series of gentamycin and kanamycin in cation-adjusted Mueller Hinton Broth (MH) (Oxoid) were set up in microtiter plates with final concentrations ranging from 256 to 4 mg/L. Four to ten colonies from each isolate were homogenized in 0.9% NaCl to achieve a final concentration equivalent to 0.5 McFarland. The bacterial suspensions were further diluted 1:150 in MH before 50 μl was transferred to the microtiter wells containing 50 μl MH with gentamycin or kanamycin. The assay was further modified to suit slow growing environmental bacteria. The temperature was reduced from 35°C to 28°C, and the incubation time was increased from 16–20 h to 96 h. Growth was registered visually every day for 4 days. Escherichia coli strain ATCC 25922 was used as a quality control reference for determination of QC breakpoints as recommended by CLSI (2017) .
3
Microbial Cultivation and Nematode Propagation
4
Culturing and Storing Lactobacillus and Campylobacter
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L. johnsonii strain FI10058 is derived from strain FI9785 and contains plasmid pFI2431, produced by the insertion of a chloramphenicol resistance gene into the highly stable native small plasmid p9785 (Horn et al., 2005) . It was routinely grown at 37°C on de Man, Rogosa, Sharpe agar or broth (MRS, Oxoid) supplemented with 10 µg/ml neomycin and 7.5 µg/ml chloramphenicol. Overnight cultures used for inoculation were harvested by centrifugation at 3000 g for 15 min at 20°C, washed twice and resuspended in phosphate-buffered saline (PBS) at approximately 1×10 9 colony forming units (cfu)/ml. C. jejuni strain 81-176 was isolated from an outbreak associated with unpasteurised milk (Korlath et al., 1985) . It is known that this strain readily colonises chickens (Guccione et al., 2008) . It was routinely cultured on sheep blood agar (Oxoid) under standard microaerobic conditions (10% O 2 , 5%
CO 2 and 85% N 2 ) at 37 o C in a MACS-VA500 incubator (Don Whitley Scientific). Cultures used for inoculation were incubated for 24 h in 10 ml Mueller-Hinton broth (MH, Oxoid).
Long-term storage of Campylobacter was at -80°C in Microbank vials (Prolab Diagnostics).
Campylobacter blood-free selective agar (CBF) plates were prepared according to the manufacturer's instructions from Campylobacter blood-free selective agar (CCDA, Oxoid)
and CCDA selective supplement (Oxoid).
CO 2 and 85% N 2 ) at 37 o C in a MACS-VA500 incubator (Don Whitley Scientific). Cultures used for inoculation were incubated for 24 h in 10 ml Mueller-Hinton broth (MH, Oxoid).
Long-term storage of Campylobacter was at -80°C in Microbank vials (Prolab Diagnostics).
Campylobacter blood-free selective agar (CBF) plates were prepared according to the manufacturer's instructions from Campylobacter blood-free selective agar (CCDA, Oxoid)
and CCDA selective supplement (Oxoid).
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