L 1 yeast extract

Manufactured by Thermo Fisher Scientific

Sourced in United Kingdom

L-1 yeast extract is a high-quality nutrient source for microbial cultures. It provides a rich source of amino acids, vitamins, and other growth factors to support the cultivation of a variety of microorganisms, including yeast and bacteria.

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Lab products found in correlation

Ampicillin, by Merck Group (2 mentions) L 1 tryptone, by Thermo Fisher Scientific (2 mentions) Hemin, by Thermo Fisher Scientific (1 mentions) Horse serum, by Thermo Fisher Scientific (1 mentions) Menadione, by Merck Group (1 mentions) Anaerojar, by Thermo Fisher Scientific (1 mentions) Hemin, by Merck Group (1 mentions) L cysteine, by Thermo Fisher Scientific (1 mentions) L 1 l cysteine, by Merck Group (1 mentions) Trypticase soy agar, by Thermo Fisher Scientific (1 mentions) Luria bertani agar lb, by Merck Group (1 mentions) L 1 hemin, by Merck Group (1 mentions) Gaspak system, by Mitsubishi (1 mentions) L 1 agar, by Merck Group (1 mentions)

7 protocols using l 1 yeast extract

Cultivation of Tannerella forsythia

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Tannerella forsythia ATCC 43037 (American Type Culture Collection, USA) and defined T9SS mutants (see below) were grown in 37 g L⁻¹ of Brain-Heart-Infusion (BHI) liquid media (Oxoid, UK), containing 5 g L⁻¹ yeast extract (Oxoid), 0.5 g L⁻¹ L-cysteine (Sigma, Austria), 2.5 μg mL⁻¹ hemin (Sigma), 2.0 μg mL⁻¹ menadione (Sigma), 10 μg mL⁻¹N-acetylmuramic acid (Carbosynth, UK) and 5% (v/v) horse serum (Life Technologies, Austria), under anaerobic conditions at 37°C for 4-7 days. For cultivation of T. forsythia wild-type and mutants on BHI agar plates (0.8% w/v), the amounts of L-cysteine, hemin, and N-acetylmuramic acid were doubled and plates were incubated under anaerobic conditions in an anaerobic jar (AnaeroJar; Oxoid) at 37°C. Media were supplemented with gentamycin and erythromycin at a concentration of 200 μg mL⁻¹ and 5 μg mL⁻¹, respectively, when appropriate.
Escherichia coli strains were grown under standard conditions in Luria-Bertani (LB) medium supplemented with 100 μg mL⁻¹ ampicillin, when appropriate. P. gingivalis W83 is used as a reference strain for comparison with predicted components of the T9SS in T. forsythia ATCC 43037.

Tomek M.B., Neumann L., Nimeth I., Koerdt A., Andesner P., Messner P., Mach L., Potempa J.S, & Schäffer C. (2014). The S-layer proteins of Tannerella forsythia are secreted via a type IX secretion system that is decoupled from protein O-glycosylation. Molecular oral microbiology, 29(6), 307-320.

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Cultivating Saccharomyces cerevisiae in µBR

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Saccharomyces cerevisiae (YSC2, Sigma‐Aldrich, Dorset, UK) inoculum was prepared from mid‐exponential YPD10 medium shake flask cultures. The YPD10 medium consisted of 10 g L ⁻¹ yeast extract (Oxoid Ltd, UK), 20 g L⁻¹ Bacto™ Peptone (BD Biosciences, UK), 10 g L⁻¹ D‐glucose (Fisher Scientific, UK) and 50 mg L⁻¹ ampicillin (Sigma‐Aldrich, UK). Growth was performed in 500 mL baffled shaken flasks, with a headspace of 80%, with 50 mm shaking diameter, at a shaking frequency of 200 rpm and 30°C.
The µBR was sterilized by priming with 70% (v/v) ethanol for 30 min (Supporting Information 2), followed by three cycles of flushing/priming with autoclaved deionized (DI) water at intervals of 20 min each. The µBR was filled with YPD10 medium and inoculated with S. cerevisiae. The inoculum syringe was then exchanged with a syringe system containing DI water. Mixing was achieved by withdrawing and infusing a nominal volume of 150 µL with a syringe drive pump (World Precision Instruments, Inc., USA). The pump was operated at a flow rate of 3 mL min⁻¹ during withdrawal and 5 mLmin⁻¹ during infusion, with a full cycle taking approximately 4 s. End‐point OD values were measured to determine average cell concentration in the µBR. Due to the small volumes involved, the dry weight was determined via the correlation proposed by Oh et al.18

Kirk T.V., Marques M.P., Radhakrishnan A.N, & Szita N. (2016). Quantification of the oxygen uptake rate in a dissolved oxygen controlled oscillating jet‐driven microbioreactor. Journal of Chemical Technology and Biotechnology, 91(3), 823-831.

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Salmonella Typhimurium Genetic Manipulation

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All experiments were performed with Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 (designated S. enterica) and derivatives thereof. Mutations were transferred between strains through generalized transduction with phage P22 HT 105/1 int-201 (Schmieger, 1972 (link)). All growth was done in LB [5 g L^–1 yeast extract (Oxoid), 10 g L^–1 Tryptone (Oxoid), 10 g L^–1 NaCl, and 1 mM NaOH] or LA (LB solidified with 15 g L^–1 agar). To simplify preparation of competent cells for electroporation, NaCl was omitted from the LB medium. When needed, antibiotics were added to the following concentrations: chloramphenicol (cam); 12.5 mg L^–1, ampicillin (amp); 50 or 100 mg L^–1, kanamycin (kan); 50 mg L^–1, and tetracycline (tet); 7.5 mg L^–1. To select for loss of the cat-sacB cassette, we used sucrose selection plates (LA without NaCl, supplemented with 50 g L^–1 sucrose). When growing cells to exponential phase to prepare samples for LC-MS/MS, FACS analysis or mRNA extractions for RT-PCR, LB was supplemented with 2 g L^–1 glucose in order to allow for exponential growth at higher cell density. All growth (except during λ red recombineering) was at 37°C.

Knöppel A., Andersson D.I, & Näsvall J. (2020). Synonymous Mutations in rpsT Lead to Ribosomal Assembly Defects That Can Be Compensated by Mutations in fis and rpoA. Frontiers in Microbiology, 11, 340.

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Anaerobic Growth of T. forsythia 92A2

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T. forsythia 92A2 was grown anaerobically using a GasPak system (Mitsubishi Gas Chemical Company Inc) at 37°C for 3–7 days on Trypticase Soy Agar (TSA) supplemented with 5 g L⁻¹ yeast extract (Oxoid), 1 g L⁻¹ L-cysteine, 10 mg L⁻¹ N-acetylmuramic acid, 5 mg L⁻¹ hemin, 0.5 mg L⁻¹ menadione (all reagents from Sigma Aldrich) and 50 ml L⁻¹ defibrinated sheep blood (Northeastern Lab). Liquid cultures were done in Trypticase Soy Broth (TSB) supplemented as noted above without sheep blood. Escherichia coli BL21DE3 was grown on Luria-Bertani agar (LB) (Sigma Aldrich) overnight aerobically at 37°C. After pET-22b⁺ plasmid cloning, E.coli BL21DE3 (Novagen) was grown on LB agar was supplemented with 50 mg ml⁻¹ of ampicillin (Sigma Aldrich).

Yost S, & Duran-Pinedo A.E. (2018). The contribution of Tannerella forsythia dipeptidyl aminopeptidase IV in the breakdown of collagen. Molecular oral microbiology, 33(6), 407-419.

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Lead Biosorption by Bacteria

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The batch reactors and agar plates contained Luria–Bertani (LB) broth consisting of 1 g L

^{- 1}

NaCl (Glassworld, South Africa), 20 g L

^{- 1}

tryptone (Oxoid, UK) and 10 g L

^{- 1}

yeast extract (Oxoid, UK), with 5 g L

^{- 1}

agar (Sigma Life Science, Spain) added to the agar plates solution only. The stock solution of 10,000 mg L

^{- 1}

Pb(NO

_{3}

)

_{2}

consisted of 1.6 mg Pb(NO

_{3}

)

_{2}

(Glassworld, South Africa) in 100 mL distilled water. Metabolic activity was determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) (Sigma, Aldrich, MO, USA). The nitrate concentration was determined using a nitrate test (Supelco, Germany). Ethylenedinitrilotetraacetic acid disodium salt (EDTA) (Supelco, Germany) was used in the determination of the extracellular and intracellular Pb(II) concentration.

Neveling O., Ncube T.M., Ngxongo Z.P., Chirwa E.M, & Brink H.G. (2022). Microbial Precipitation of Pb(II) with Wild Strains of Paraclostridium bifermentans and Klebsiella pneumoniae Isolated from an Industrially Obtained Microbial Consortium. International Journal of Molecular Sciences, 23(20), 12255.

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Anaerobic Growth Analysis of Microbes

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For in vitro growth analysis in a 200-μl micro-volume a Bioscreen C growth curve reader was used (Oy Growth Curves Ab Ltd., Helsinki, Finland). Overnight cultures were diluted 1:200 in a modified minimal medium supplied with 0 or 10 mM sodium nitrate (NaNO₃). The composition of the medium with glutamine as nitrogen source was described previously (Kaspar et al., 2014 (link)). As carbon source 1 g L⁻¹ D (+)-glucose monohydrate (Fluka, Darmstadt, Germany) was used. Furthermore, 5 g L⁻¹ yeast extract (Oxoid, Wesel, Germany) were added. Cultures were incubated at 34°C with continuous medium shaking (shaking step 60). For anaerobic growth analyses, cultures were overlaid with 200 μl sterile paraffin oil (Roth, Karlsruhe, Germany). The OD₆₀₀ of each well was automatically recorded every hour over a period of 24 h. Mean values and standard deviation were calculated from at least four independent biological experiments each including technical duplicates. A student's t-test (Two-Sample Assuming Unequal Variance, Microsoft Excel) was performed for statistical analysis.

Ferrari E., Walter M.C., Huptas C., Scherer S, & Müller-Herbst S. (2017). Complete Circular Genome Sequence and Temperature Independent Adaptation to Anaerobiosis of Listeria weihenstephanensis DSM 24698. Frontiers in Microbiology, 8, 1672.

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Bacterial Growth Conditions and Genetic Manipulations

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The strains and plasmids used in this study are described in S1 Table and S2 Table. Bacterial cultures were routinely grown in lysogeny broth (LB) broth at 37°C with aeration or on LB solidified with 1.5% agar (BD Biosciences, San Jose, CA). When stated, bacteria were grown in PS:DB media, which consists of development buffer (DB) (5 mM KH₂PO₄, 5 mM Na₂HPO_4, 2 mM MgCl₂, 1 mM CaCl₂ pH 6.5) and 10% (v/v) PS medium (10 g L^-1 Special Peptone (Oxoid, Hampshire, United Kingdom), 7 g L^-1 Yeast Extract (Oxoid, Hampshire, United Kindom), 10 mM KH₂PO₄, 0.45 mM Na₂HPO₄, 15 g L^-1 glucose, 20 nM vitamin B12, 180 nM Folic Acid, pH 6.5). Antibiotics were added at the following concentrations when appropriate: carbenicillin 300 μg mL^-1, gentamycin 30 μg mL^-1, and tetracycline 200 μg mL^-1 for P. aeruginosa; 100 μg mL^-1, gentamycin 30 μg mL^-1, and tetracycline 15 μg mL^-1 for E. coli. Expression of P_tac- or P_lac-controlled genes was induced with 1 mM IPTG. When indicated, cultures were supplemented with HHQ, PQS, or HQNO (Cayman Chemicals, Ann Arbor, MI). Unless otherwise stated, chemicals and reagents were purchased from Sigma Aldrich (St. Louis, MO).

Vrla G.D., Esposito M., Zhang C., Kang Y., Seyedsayamdost M.R, & Gitai Z. (2020). Cytotoxic alkyl-quinolones mediate surface-induced virulence in Pseudomonas aeruginosa. PLoS Pathogens, 16(9), e1008867.

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