The fermentation capacities were analyzed quantitatively by ethanol determination. One isolated strain (Mi4) was inoculated in culture media (3.0 g/L of yeast extract (Difco Laboratories, Detroit, MI, USA) and 5.0 g/L of proteose peptone No. 3 (Difco Laboratories, Detroit, MI, USA) with different carbon sources: galactose (1%) or lactate (1%) plus galactose (1%) or sucrose (1%) (Sigma Aldrich Chemie, Steinheim, Germany). After seven days the ethanol produced was measured with Enzytec fluid Ethanol purchased from R-Biopharm, Darmstadt, Germany (Cat. No. E5340), following the instructions supplied by the manufacturer.
Proteose peptone no 3
Manufactured by BD
Sourced in Germany, United States
Proteose Peptone No. 3 is a product designed for use in microbiological culture media. It serves as a source of nitrogen, amino acids, and other nutrients to support the growth of a variety of microorganisms in laboratory settings.
Automatically generated - may contain errors
Lab products found in correlation
Soluble starch, by Merck Group (7 mentions)
Sodium chloride (nacl), by Carl Roth (6 mentions)
Thiamine pyrophosphate, by Merck Group (6 mentions)
D glucose, by Carl Roth (6 mentions)
L glutamine, by Carl Roth (6 mentions)
4 aminobenzoic acid, by Merck Group (6 mentions)
Vitamin b12, by Merck Group (6 mentions)
K2hpo4, by Carl Roth (5 mentions)
Kh2po4, by Carl Roth (5 mentions)
Thiamine hcl, by Carl Roth (5 mentions)
Fe no3 3, by Merck Group (5 mentions)
Dehydrated agar, by BD (4 mentions)
β nicotinamide adenine dinucleotide, by Carl Roth (4 mentions)
Yeast extract, by BD (3 mentions)
Chloramphenicol, by Merck Group (3 mentions)
Casamino acid, by BD (3 mentions)
Yeast extract, by Thermo Fisher Scientific (3 mentions)
Sodium pyruvate, by Merck Group (3 mentions)
Bacto agar, by BD (3 mentions)
Magnesium sulfate heptahydrate, by Merck Group (2 mentions)
18 protocols using proteose peptone no 3
1
Quantifying Ethanol Production from Diverse Sugars
2
Isolation and Cultivation of Yeast Strains
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All strains isolated in this study come from the same industry and are listed in Table 1 . In addition, seven reference strains of M. guilliermondii from the Spanish Type Culture Collection (CECT) were included for comparison in the typing study (Table 1 ), as well as a Saccharomyces cerevisiae strain as fermentation control. The culture media used were YMB (Yeast morphology broth), YMA (Yeast morphology agar), and YMAC (YMA plus Chloramphenicol). YMB had 10.0 g/L glucose (Panreac Química, Barcelona, Spain), 5.0 g/L proteose peptone No. 3 (Difco Laboratories, Detroit, MI, USA), 3.0 g/L yeast extract (Difco), and 3.0 g/L malt extract (Difco). The YMA was YMB solidified with 20.0 g/L agar. YMAC was made by adding 0.5 g/L of Chloramphenicol (Sigma Aldrich Chemie, Steinheim, Germany) to YMA. To isolate the strains, 10 g of the samples were suspended in YMB, homogenized in a Stomacher homogenizer, and serial dilutions in saline solution were made. To enumerate the viable cells, two replicas with four drops (50 μL) of an appropriate dilution were inoculated on YMAC, following the modified method of Miles and Misra [11 (link),12 ]. Strains were routinely grown at 28 °C in YMB and maintained on YMA slants at 4 °C.
3
Cultivation of Gardnerella species
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G. vaginalis ATCC 14018T, G. leopoldii UGent 06.41T, G. piotii UGent 18.01T and G. swidsinskii GS 10234T were grown on chocolate (Choc) agar plates (Becton Dickinson, Franklin Lakes, NJ, USA) at 37 °C under anaerobic conditions (10% CO2, 10% H2, 80%N2, Concept 400, Anaerobic Workstation). Liquid cultures were prepared in New York City Broth III (10 mM HEPES (Sigma Aldrich, Burlington, MA, USA), 15 g/L Proteose Peptone No. 3 (Becton Dickinson), 3.8 g/L yeast extract (Thermo Fisher Scientific, Waltham, MA, USA), 86 mM sodium chloride (Carl Roth, Karlsruhe, Germany), 28 mM α-D-glucose (Sigma–Aldrich)), at pH 5 (NYB5), supplemented with 10% horse serum (HS, Thermo Fisher Scientific) (NYB5 + HS).
4
Cultivation and Characterization of C. perfringens
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C. perfringens CCRC 10,648 and CCRC 13,019 were purchased from the Biosources Collection and Research Center (Hsinchu, Taiwan, China). Acetic acid, acetonitrile, dimethyl sulfoxide (DMSO), glycerol, methanol, and sodium hydroxide (NaOH) were purchased from Fluka (Garage Gmbh, Buchs, Switzerland). Meanwhile, sodium azide (NaN3), phenol red, and sodium bicarbonate (NaHCO3) were purchased from Sigma Chemical Co. (Gillingham, UK). Cellulase (3000 U/g) from Trichoderma viride was purchased from Challenge Bioproducts Co., Ltd. (Taichung, Taiwan, China), whereas chitin powder was obtained from Applied Chemical Co., Ltd. (Kaohsiung, Taiwan, China). LIB, plate count agar (PCA), tryptose–sulfite–cycloserine (TSC) agar base, bacto agar, proteose peptone no. 3, yeast extract, and 50% egg yolk saline were supplied by Becton Dickinson (Sparks, MD, USA).
5
Cultivation Protocols for Diverse Bacterial Isolates
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Unless stated otherwise, At-SPHERE isolates were
cultivated at room temperature on R-2A agar (Sigma-Aldrich) or at 28°C in
R-2A broth (0.5 g yeast extract (Oxoid), 0.5 g Proteose peptone No. 3 (Becton,
Dickinson and Company), 0.5 g Casamino acids (Becton, Dickinson and Company),
0.5 g D-glucose monohydrate (Sigma-Aldrich), 0.5 g starch (from potato, Fluka),
0.3 g sodium pyruvate (Sigma-Aldrich), 0.3 g K2HPO4(AppliChem) and 0.05 g magnesium sulfate heptahydrate (Sigma-Aldrich) dissolved
in 1 L deionized water), both supplemented after sterilization with 0.5% (v/v)
methanol (R-2A+M). Minimal medium was prepared as described previously66 (link) and supplemented with 5 mM
maltose (Fluka). For Leaf374 cultivation, minimal medium agar was supplemented
with vitamins (500 μg L-1 D-pantothenic acid hemi calcium
salt, 100 μg L-1 biotin, 400 μg L-1riboflavin, 400 μg L-1 thiamine HCl, 200 μg
L-1 pyridoxal HCl 150 μg L-1 p-amino benzoic
acid, 200 μg L-1 cobalamin, 50 μg L-1 lipoic
acid, 150 μg L-1 nicotinic acid and 100 μg
L-1 folic acid). For selective growth of Leaf272, R-2A+M was
supplemented with 50 μg mL-1 rifampicin (Sigma-Aldrich), since
this strain has a natural resistance towards this antibiotic.
E. coli BL21 (DE3) gold was cultivated in LB-Lennox at
37°C supplemented with 50 μg mL-1 kanamycin sulfate
(AppliChem).
cultivated at room temperature on R-2A agar (Sigma-Aldrich) or at 28°C in
R-2A broth (0.5 g yeast extract (Oxoid), 0.5 g Proteose peptone No. 3 (Becton,
Dickinson and Company), 0.5 g Casamino acids (Becton, Dickinson and Company),
0.5 g D-glucose monohydrate (Sigma-Aldrich), 0.5 g starch (from potato, Fluka),
0.3 g sodium pyruvate (Sigma-Aldrich), 0.3 g K2HPO4(AppliChem) and 0.05 g magnesium sulfate heptahydrate (Sigma-Aldrich) dissolved
in 1 L deionized water), both supplemented after sterilization with 0.5% (v/v)
methanol (R-2A+M). Minimal medium was prepared as described previously66 (link) and supplemented with 5 mM
maltose (Fluka). For Leaf374 cultivation, minimal medium agar was supplemented
with vitamins (500 μg L-1 D-pantothenic acid hemi calcium
salt, 100 μg L-1 biotin, 400 μg L-1riboflavin, 400 μg L-1 thiamine HCl, 200 μg
L-1 pyridoxal HCl 150 μg L-1 p-amino benzoic
acid, 200 μg L-1 cobalamin, 50 μg L-1 lipoic
acid, 150 μg L-1 nicotinic acid and 100 μg
L-1 folic acid). For selective growth of Leaf272, R-2A+M was
supplemented with 50 μg mL-1 rifampicin (Sigma-Aldrich), since
this strain has a natural resistance towards this antibiotic.
E. coli BL21 (DE3) gold was cultivated in LB-Lennox at
37°C supplemented with 50 μg mL-1 kanamycin sulfate
(AppliChem).
6
Cultivation Protocols for Diverse Bacterial Isolates
7
Bacterial Growth in Mercury-Contaminated Media
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Bacteria were grown in modified King's B medium [KB: 10 g glycerol, 1.5 g MgSO4.7H2O, 1.5 g K2HPO4.3H2O, 20 g Proteose peptone No. 3 (BD Company) l−1], supplemented with antibiotics (30 μg ml−1 gentamicin or 250 μg ml−1 streptomycin), 10–60 μM HgCl2 [Hg(II)] and 1.2% w/v agar, where required. Inocula for experiments were provided by 18 h overnight shaken cultures incubated at 28°C. Loosely lidded Universal bottles (30 ml) were used for microcosms: KB microcosms contained 6 ml KB, soil microcosms contained 10 g of twice‐autoclaved John Innes #2 potting soil with ∼ 25% w/v water content and were mixed with 800 μl 0–4 mM Hg(II) and allowed to equilibrate for 1 h before use.
8
Gonococcal Base Agar Preparation
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Media composition, assay preparation, and setup follow [27 (link)]. Gonococcal base agar was made from 10 g/L dehydrated agar (BD Biosciences, Bedford, Massachusetts, United States of America), 5 g/L NaCl (Roth, Darmstadt, Germany), 4 g/L K2HPO4 (Roth), 1 g/L KH2PO4 (Roth), 15 g/L Proteose Peptone No. 3 (BD Biosciences), 0.5 g/L soluble starch (Sigma-Aldrich, St. Louis, MO), and supplemented with 1% IsoVitaleX (IVX): 1 g/L D-glucose (Roth), 0.1 g/L L-glutamine (Roth), 0.289 g/L L-cysteine-HCL × H2O (Roth), 1 mg/L thiamine pyrophosphate (Sigma-Aldrich), 0.2 mg/LFe(NO3) 3 (Sigma-Aldrich), 0.03 mg/L thiamine HCl (Roth), 0.13 mg/L 4-aminobenzoic acid (Sigma-Aldrich), 2.5 mg/L β-nicotinamide adenine dinucleotide (Roth), and 0.1 mg/L vitamin B12 (Sigma-Aldrich). GC medium is identical to the base agar composition but lacks agar and starch. Employed bacterial strains (Table i in S1 Text ) are derivatives of strain N. gonorrhoeae MS11.
9
Bacterial Growth Across Temperatures
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Bacteria were cultured at 20, 28, 37, and 42°C using Brain Heart Infusion (BHI; Oxoid, UK), King's B (KB; 20 g proteose peptone No. 3 (BD Biosciences, UK), 10 g glycerol, 5 g K 2 HPO 4 , 1.5 g MgSO 4 per litre) and Luria-Bertani (LB; 10 g NaCl, 10 g
10
Isolation and Analysis of Neutrophils
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