Fifteen strains belonging to 12 dominant and subdominant butyrate-producing species from the Lachnospiraceae and Ruminococcaceae families with publicly available genomes were assessed for vitamin and amino acid auxotrophies (Fig. 1 ; see also Table S3 in the supplemental material). Stocks were routinely kept in M2GSC medium (35 (link)) and subcultured twice in chemically defined medium (CDM) (see below) before the start of the experiments (1% inoculum). Bifidobacterium bifidum CNCM I-3650, Lactobacillus paracasei CNCM I-1518, and Streptococcus thermophilus CNCM I-3862 (Table S3 ) from the Danone strain collection were used for coculture experiments. They were maintained in medium M17 plus 1 g/liter l- cysteine (S. thermophilus CNCM I-3862) or MRS plus 1 g/liter l- cysteine (B. bifidum CNCM I-3650, L. paracasei CNCM I-1518) (media were purchased from Oxoid, Thermo Fisher Scientific, Waltham, MA, USA) and subcultured in CDM before coculture experiments.
L cysteine
Manufactured by Thermo Fisher Scientific
Sourced in United States, United Kingdom
L-cysteine is a non-essential amino acid that is commonly used in various laboratory applications. It serves as a building block for proteins and plays a role in the production of other biomolecules. L-cysteine can be used in a range of research and analytical procedures, but a detailed, unbiased, and non-interpretative description of its core function is not available.
Automatically generated - may contain errors
Lab products found in correlation
Pd 10 desalting column, by GE Healthcare (7 mentions)
Ferric chloride, by Thermo Fisher Scientific (7 mentions)
Sodium sulfide, by Thermo Fisher Scientific (6 mentions)
Dithiothreitol (dtt), by Thermo Fisher Scientific (5 mentions)
Hemin, by Merck Group (4 mentions)
L methionine, by Merck Group (4 mentions)
Yeast extract, by Thermo Fisher Scientific (4 mentions)
Cefoxitin, by Thermo Fisher Scientific (4 mentions)
L glutamine, by Thermo Fisher Scientific (3 mentions)
Dnase 1, by Roche (3 mentions)
Vitamin k1, by Merck Group (3 mentions)
Dulbecco modified eagle medium (dmem), by Merck Group (3 mentions)
Dl homocysteine, by Merck Group (3 mentions)
Sodium chloride (nacl), by Merck Group (3 mentions)
Tryptic soy broth (tsb), by BD (3 mentions)
Nupage lds sample buffer, by Thermo Fisher Scientific (2 mentions)
High glucose dmem, by Thermo Fisher Scientific (2 mentions)
L methionine, by Thermo Fisher Scientific (2 mentions)
Anti ha agarose beads, by Merck Group (2 mentions)
35s methionine cysteine, by PerkinElmer (2 mentions)
59 protocols using l cysteine
1
Assessing Auxotrophies of Butyrate-Producers
2
Radiolabeling Shank3 Dynamics
3
Porcine Oocyte Maturation Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All chemicals and reagents were purchased from Sigma-Aldrich (Oslo, Norway) unless otherwise stated. Porcine X medium (PXM) was used for washing cumulus-oocyte complexes (COCs) and porcine oocyte medium (POM) for oocyte maturation [25 (link)] with minor modifications to the POM (20 ml/l BME amino acids, 10.0 ml/l MEM non-essential amino acids, 5.0 mM glucose, 0.2 mM Na-pyruvate, 0.6 mM L-cysteine, 2.0 mM Ca-(lactate)2·5H2O, 2.0 mM l -glutamine, 108 mM NaCl, 10 mM KCl, 25 mM NaHCO3, 0.4 mM MgSO4·7H2O, 0.35 mM KH2PO4, 5.0 mM hypotaurine, 50 µM β-mercaptoethanol (Gibco, Fisher Scientific AS, Oslo, Norway), 10 ng/ml epidermal growth factor, 0.01 mg/ml gentamicin, FLI (FGF2 40 ng/mL, LIF 20 ng/mL, IGF1 20 ng/mL) and 4.0 mg/ml BSA).
4
Radiolabeling Shank3 Dynamics
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HeLa cells cultured in 6-well plates were transfected with 500 ng plasmids expressing HA-Shank3 (WT or 3A) by Lipofectamine 3000. 30 hr later, methionine starvation was performed for 1hr by changing the culture media with 1 mL pulse medium (DMEM high glucose without L-glutamine, sodium pyruvate, L-methionine, and L-cysteine [Gibco] + 10% FBS + 2 mM L-glutamine) into each well. After methionine starvation, pulse medium was changed by labeled pulse medium (pulse medium plus 100 μCi 35S methionine/cysteine [PerkinElmer]) and cells were incubated for 30min. After labeling, cells were washed with warm PBS for 3 times and then 1 mL chase medium (DMEM + 10% FBS + 2mM L-methionine) was added into each well. Cells were harvested at indicated time points after adding chase medium. Harvested cells were lysed in 1 mL DOC buffer (50 mM Tris-HCl pH 9.0, 5mM EDTA, 1% sodium deoxycholate) supplemented with phosphatase and protease inhibitors (GenDEPOT) and incubated with anti-HA-agarose beads (Sigma) for 2 hr at 4 °C. The beads were washed with DOC buffer for 3 times, boiled with NuPAGE LDS sample buffer (Invitrogen) to elute proteins. The eluted products were run on an SDS-PAGE gel. The gel was fixed for 30 min in a 2% salicylic acid, 30% methanol solution and dried before detection by autoradiography.
5
Dissociation and Culture of Electroporated Cortical Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Pair-cell analysis was performed as described (Shen et al., 2002 (link)). Following electroporation at E14.5 as described above, electroporated brains at E16.5 and E17.5 were sectioned using a vibratome (Leica, VT1000S). Cortical tissues that were enriched for transferred cells (marked by GFP) were dissected with tungsten needles from cortical sections. For the cortex transfected with Mek1DD, the upper and lower half of the cortex were further separated. To dissociate cells, dissected tissues were incubated in a protease solution containing 10 unit/ml papain (Fluka, Japan), 1000 units/ml DNAse I (Roche, Switzerland) and 5 mM L-cysteine in DMEM (Invitrogen), and triturated using a fire-polished Pasteur pipette to create a single-cell suspension. Cells were resuspended in culture medium containing DMEM, glutamine, penicillin/streptomycin, sodium pyruvate (Invitrogen), 1 mM N-acetyl-L-cysteine (Sigma), B27, N2 and 10 ng/ml bFGF2 (Invitrogen) and plated onto coverslips coated with poly-L-lysine (Sigma) at clonal density. The cultures were maintained in a humidified incubator at 37°C with constant 5% CO2 supply. In 24 or 48 hr later, the cultures were fixed and immunostained for GFP, Fabp7, and Tubb3 and counterstained with the DNA dye Hoechst 33342 solution (Invitrogen).
6
Endothelial Cell Culture Protocols
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All culture media (except EBM-2 and EGM-2), L-cysteine, fibronectin, trypsin, penicillin, and streptomycin were from Invitrogen (Breda, the Netherlands). EBM-2 and EGM-2 were from Lonza (Verviers, Belgium). Epinephrine, thrombin, forskolin, 3-isobutyl-1-methylxanthine (IBMX), LY294002, Y27632, BAPTA-AM, Endothelial Cell Growth Supplement (ECGS), and anti-α-tubulin monoclonal antibody (DM1A) were from Sigma-Aldrich Chemie (Steinheim, Germany). S1P was from Avanti Polar Lipids (Alabaster, Alabama, USA). Anti-β-catenin rabbit polyclonal antibody (sc-7199) was from Santa Cruz Biotechnology (Santa Cruz, California, USA). Chemiluminescence blotting substrate and Complete Protease Inhibitor Cocktail Tablets were from Roche Diagnostics (Mannheim, Germany). All chemicals used were of analytical grade. Anti-VWF monoclonal antibody CLB-RAg20 has been described previously [29] (link). Enzyme-linked immunosorbent assays (ELISA) for VWF and VWF propeptide have been described previously [30] (link).
7
Cultivation of Faecalibacterium duncaniae
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Faecalibacterium duncaniae DSM 17677 strain (Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany) was used in this study. It was initially cultured according to the recommended conditions proposed by DSMZ [21 ]. For long term storage, this strain was kept frozen at −80 °C in sBHI broth [Brain Heart Infusion medium (37 g/L; VWR International, Leuven, Belgium) supplemented with yeast extract (5 g/L; VWR International), hemin (5 mg/L; Alfa Aesar, Kandel, Germany), vitamin K1 (5 µL/L; Sigma-Aldrich Co., St. Louis, MO, USA), and L-cysteine (2 g/L; Alfa Aesar)], as previously used by Maier et al. [22 (link)], with 20% (v/v) of glycerol (Fisher Scientific, Loughborough, UK). For each assay, a F. duncaniae glycerol stock was thawed and grown in sBHI broth for 16 h at 37 °C under anaerobic conditions (85% N2, 5% H2, and 10% CO2) achieved in an anaerobic incubator (Whitley A35 HEPA anaerobic workstation, Bingley, UK). Afterwards, the previously grown culture was transferred to sBHI broth (in a proportion of 1:100), and this bacterial suspension was anaerobically incubated during 10 h at 37°C for the following experiments.
8
Inducing ER Stress in Breast Cancer Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MB468 and MB468res-R8 cell lines were maintained in DMEM (Sigma-Aldrich, D0422) supplemented with 10% dialyzed FBS (Omega Scientific), 1.5 μM cyanocobalamin (vitamin B12), 4 mM L-glutamine, 100 μM L-cysteine (Fisher Scientific), and 100 μM L-methionine (Sigma-Aldrich). In the case of methionine-free media, 370 μM DL-homocysteine (Sigma-Aldrich) was added in the absence of methionine.
To induce ER stress, MB468 and MB468res-R8 cells were treated with the 1 μM thapsigargin (Sigma-Aldrich, T9033) for 4 h. To inhibit PERK activation, cells were treated with 1 μM GSK2656157 (Millipore Sigma, 5.04651.0001) 1 h before media switch or thapsigargin treatment and replaced in the new media.
To induce ER stress, MB468 and MB468res-R8 cells were treated with the 1 μM thapsigargin (Sigma-Aldrich, T9033) for 4 h. To inhibit PERK activation, cells were treated with 1 μM GSK2656157 (Millipore Sigma, 5.04651.0001) 1 h before media switch or thapsigargin treatment and replaced in the new media.
9
Prostate Cancer Cell Line Culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human normal prostate stromal immortalized cell line WPMY-1 (BNCC100291) and PCa cell lines PC-3 (BNCC337715), Du 145 (BNCC338240), LNCaP (BNCC337703), and 22RV1 (BNCC100161) were bought from BeNa Culture Collection (Shanghai, China). Cells were prepared at 37℃ with 5% CO2. Culture medium information was: PC-3, Du 145, WPMY-1 cell lines in DMEM-H (BNCC338068) plus 10% FBS; LNCaP, 22RV1 cell lines in RPMI-1640 plus 10% FBS (BNCC341471). Du 145 cells were passaged in DMEM (Sigma-Aldrich) plus 10% dialyzed FBS (Gemini Bio-Products) in the methionine dependence assay. Control medium was supplemented with 100 µM l -methionine (Sigma-Aldrich), 100 µM l -cysteine (Fisher Scientific), 1.5 µM vitamin B12, and 4 mM l -glutamine. Also, 200 µM dl -homocysteine (Sigma-Aldrich) was added to the Met-Hcy+ medium. For sulfasalazine treatment, sulfasalazine (HY-14655, medchemexpress, USA) was purchased and dissolved in DMSO, and was added to the culture medium at a final concentration of 200 µM for treating PCa cells. The same amount of DMSO was supplemented to the control group for treatment [33 (link)].
10
Isogenic Francisella iglD Mutant Strains
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The Fn iglD strain KKF37 [19] (link) is isogenic with wildtype Fn strain U112 and the Ftt iglD strain KKT8 is isogenic with wildtype Ftt strain Schu S4. The Ftt iglD strain had both copies of iglD (iglD1 iglD2) inactivated by a Group II intron targeted to iglD, as described in [35] (link). Francisella strains were grown in tryptic soy broth (TSB) (BD Biosciences) supplemented with 0.1% (w/v) L-cysteine (Fisher Scientific) and sodium metabisulfite (Sigma), iron sulfate (Mallinckrodt), and sodium pyruvate, all at 250 µg/ml final concentration, or Chamberlain's defined medium [36] (link).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!