Knockout dulbecco s modified eagle s medium

Manufactured by Thermo Fisher Scientific

Sourced in United States, Canada

KnockOut Dulbecco's modified Eagle's medium is a cell culture medium formulated to support the growth and maintenance of embryonic stem cells and other pluripotent cell types. It is optimized to maintain the undifferentiated state of these cells.

Automatically generated - may contain errors

Lab products found in correlation

15 protocols using knockout dulbecco s modified eagle s medium

Generation of Transgenic mESC Reporter Line

Check if the same lab product or an alternative is used in the 5 most similar protocols

G4 hybrid ESCs [23 (link)] were tested negative for mycoplasma contamination and cultured in Knockout Dulbecco’s modified Eagle’s medium (Invitrogen), high-glucose, supplemented with 15% v/v fetal calf serum (Promega), 0.1 mM nonessential amino acids (Invitrogen), 2 mg/ml l-Glutamine (Invitrogen), 50 μg/ml each penicillin and streptomycin (Invitrogen), 0.1 mM β-mercaptoethanol (Sigma), and 500 U/ml leucemia inhibitory factor (Chemicon). The ESCs were kept on irradiated murine fibroblasts derived from neomycin resistant mice.
mESCs were transfected with the CAG-sA5-YFP plasmid by electroporation. After neomycin (Geneticin Sulfate, Gibco/Life Technologies) selection, resistant clones were isolated under fluorescence light to detect YFP expression. Clones with stable YFP expression were analyzed by fluorescence microscopy and qPCR. For the qPCR analysis, we used RNA isolated from wild-type G4-mESCs (provided by Nagy Lab, Toronto, Canada) [23 (link)] as negative control and RNA from an mESC line stably expressing EYFP (kindly donated by Tobias Bruegmann, Institute of Physiology I, University of Bonn, Germany) as positive control. The newly generated mESCs were karyotyped and further cultured for generation of a transgenic mouse line. Offspring were viable and did not show an overt phenotype.

Martínez-Lagunas K., Yamaguchi Y., Becker C., Geisen C., DeRuiter M.C., Miura M., Fleischmann B.K, & Hesse M. (2019). In vivo detection of programmed cell death during mouse heart development. Cell Death and Differentiation, 27(4), 1398-1414.

+ Open protocol

+ Expand

Mouse Embryonic Stem Cell Expansion

Check if the same lab product or an alternative is used in the 5 most similar protocols

R1 mouse ESCs (A. Nagy, Toronto, Canada) were expanded feeder-free on 0.1% gelatin-coated cell culture flasks (Stem Cell Technologies, Vancouver, Canada) in Growth Medium containing Knockout Dulbecco's Modified Eagle's Medium (Invitrogen, Grand Island, NY) supplemented with 15% ES-Cult Fetal Bovine Serum (Stem Cell Technologies), 1% Glutamax (Invitrogen), 1% Non-Essential Amino Acids (Invitrogen), 1% Nucleosides (Millipore, Billerica, MA), 1% Penicillin/Streptomycin (Invitrogen), 0.1 mM β-Mercaptoethanol (Invitrogen) and 10³ U/ml Leukemia Inhibitory Factor (LIF; Millipore). Medium was changed in full each day, and cells were passaged every 2 days before reaching approximately 70% confluence.

Geuss L.R., Wu D.C., Ramamoorthy D., Alford C.D, & Suggs L.J. (2014). Paramagnetic Beads and Magnetically Mediated Strain Enhance Cardiomyogenesis in Mouse Embryoid Bodies. PLoS ONE, 9(12), e113982.

+ Open protocol

+ Expand

Modulating HGPS iPSC-derived MSCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

MSCs derived from HGPS iPSCs (HGPS MSCs) and WT iPSCs (WT MSCs) were cultured in KnockOut Dulbecco’s modified Eagle’s medium (Invitrogen, Carlsbad, CA), supplemented with 20% fetal bovine serum (research grade, Sigma), 1% non-essential amino acids (Invitrogen), 1% glutamax (Invitrogen), and 0.1% β-mercaptoethanol (Invitrogen). Cell cultures were maintained at 37 °C, in 5% CO₂ in a humidified atmosphere, with the media changed every 2 days. Six hours after seeding, MSCs were treated with 0.1% dimethyl sulfoxide, 3 μM FTI (tipifarnib, R115777; Selleck Chemicals, Houston, TX), 1 μM methoctramine tetrahydrochloride (M105, Sigma), 10 μM all-trans retinoic acid (R2625, Sigma), 0.1 μM phorbol 12-myristate 13-acetate (1201, Tocris), 25 μM LY-294.002 hydrochloride (sc-215273A, Santa Cruz), 12.5 μM 8-Bromo-cAMP sodium (B7880, Sigma), 10 μM 13-cis-retinoic acid (5513, Tocris), 10 μM methotrexate (A6770, Sigma), 10 μM azathioprine (A4638, Sigma), 50 μM SB 242.084 dihydrochloride hydrate (2901, Tocris), 50 μM SMER28 (4297, Tocris). Cells were analyzed after 72 hours of treatment.

Lo Cicero A., Jaskowiak A.L., Egesipe A.L., Tournois J., Brinon B., Pitrez P.R., Ferreira L., de Sandre-Giovannoli A., Levy N, & Nissan X. (2016). A High Throughput Phenotypic Screening reveals compounds that counteract premature osteogenic differentiation of HGPS iPS-derived mesenchymal stem cells. Scientific Reports, 6, 34798.

+ Open protocol

+ Expand

Culturing hESCs on Mitomycin-C-Treated MEFs

Check if the same lab product or an alternative is used in the 5 most similar protocols

hESCs were cultured on mouse embryonic fibroblast (MEF) treated with mitomycin-C. Growth medium contained KnockOut Dulbecco's modified Eagle's medium (Gibco-Invitrogen, CA) supplemented with 15% KnockOut-SR (Gibco-Invitrogen, CA), 1mM glutamine, 0.1mM ß-mercaptoethanol (Sigma-Aldrich, MO), 1% nonessential amino acids stock (Gibco-Invitrogen, CA), penicillin (50units/ml), streptomycin (50μg/ml), and 8ng/ml FGF2 (Gibco-Invitrogen, CA). 10μM ROCK inhibitor (Y27632) was supplemented to the medium in the first 24h after thawing cells. Cells were passaged using short treatment with trypsin or trypsin-EDTA (Biological Industries, Beit Haemek, Israel) maintaining the cells as clumps of multiple cells.

Weissbein U., Plotnik O., Vershkov D, & Benvenisty N. (2017). Culture-induced recurrent epigenetic aberrations in human pluripotent stem cells. PLoS Genetics, 13(8), e1006979.

+ Open protocol

+ Expand

Culturing Haploid and Diploid hESCs

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Throughout the study we used the following cell lines: haploid hpESCs - hPES10^{26 (link)}; diploid hpESCs - SwapS4^{78 (link)} (for RNA-Seq and RRBS), pES6, pES2, pES7^{79 (link)} (for DNA methylation array); bi-parental hESCs - WA09 (H9), CSES4 (for RNA-Seq), NYSCF1, NYSCF2, HuES53, HuES64 (for DNA methylation array).
hESCs were cultured on mouse embryonic fibroblast (MEF) treated with mitomycin-C. hESC growth medium was changed every 1–2 days, containing KnockOut Dulbecco’s modified Eagle’s medium (Gibco-Invitrogen, CA) supplemented with 15% KnockOut Serum Replacement (Gibco-Invitrogen, CA), 1 mM glutamine, 0.1 mM ß-mercaptoethanol (Sigma-Aldrich, MO), 1% nonessential amino acids stock (Gibco-Invitrogen, CA), penicillin (50 U/ml), streptomycin (50 μg/ml), and 8 ng/ml FGF2 (Gibco-Invitrogen, CA). Then, 10 μM ROCK inhibitor (Y27632, Stemgent) was supplemented to the medium in the first 24 h after thawing and passaging cells. Cells were passaged using short treatment with Trypsin-EDTA (Biological Industries, Beit Haemek, Israel).

Bar S., Vershkov D., Keshet G., Lezmi E., Meller N., Yilmaz A., Yanuka O., Nissim-Rafinia M., Meshorer E., Eldar-Geva T, & Benvenisty N. (2021). Identifying regulators of parental imprinting by CRISPR/Cas9 screening in haploid human embryonic stem cells. Nature Communications, 12, 6718.

+ Open protocol

+ Expand

Murine Embryonic Stem Cell Culture

Check if the same lab product or an alternative is used in the 5 most similar protocols

Murine ES v6.5 cells were purchased from Open Biosystems. mES cells were maintained in an undifferentiated, feeder-free state in leukemia inhibitory factor (LIF) supplemented medium (Knockout Dulbecco’s modified Eagle’s medium, Invitrogen, Carlsbad, CA) with 15% ES-FBS (Invitrogen), 0.1 mM β-mercaptoethanol, 2 mM L-glutamine (Invitrogen), 0.1 mM nonessential amino acids (Invitrogen), 1000 U/mL recombinant LIF (Chemicon, Temecula CA), and 2 mM HEPES (Invitrogen). Cells were cultured on gelatin-coated (0.1% gelatin in PBS, coated for 2h at 37°C) T-75 flasks at 37C, 5% CO₂ in a humidified incubator. Cells were passaged every 2–3 days to maintain an undifferentiated state.
For initial differentiation assays, mES cells were introduced to collagen IV flasks (BD Biosciences, San Jose, CA) and maintained in α-minimum essential medium (α-MEM) (Invitrogen) supplemented with 10% ES-FBS (Invitrogen), 0.1 mM β-mercaptoethanol, 2 mM L-glutamine (Invitrogen), 0.1 mM nonessential amino acids (Invitrogen), and 2 mM HEPES (Invitrogen). Media was refreshed daily. After 5 days, cells were isolated for magnetic-activated cell sorting (MACS).

Gluck J.M., Delman C., Chyu J., MacLellan W.R., Shemin R.J, & Heydarkhan-Hagvall S. (2014). Microenvironment influences vascular differentiation of murine cardiovascular progenitor cells. Journal of biomedical materials research. Part B, Applied biomaterials, 102(8), 1730-1739.

+ Open protocol

+ Expand

Culturing Human Embryonic Stem Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Human ESCs (CSES37 (link)38 (link) and HUES14 (ref. 39 (link))) were cultured on mouse embryonic fibroblast treatment with mitomycin-C. Culture medium contained KnockOut Dulbecco's modified Eagle's medium (Gibco-Invitrogen, CA) supplemented with 15% KnockOut-SR (Gibco-Invitrogen, CA), 1 mM glutamine, 0.1 mM β-mercaptoethanol (Sigma-Aldrich, MO), 1% non-essential amino-acid stock (Gibco-Invitrogen, CA), penicillin (50 U ml⁻¹), streptomycin (50 μg ml⁻¹), and 8 ng ml⁻¹ fibroblast growth factor 2 (Gibco-Invitrogen, CA). Cells were passaged using trypsin-EDTA (Biological Industries, Beit Haemek, Israel).

Weissbein U., Schachter M., Egli D, & Benvenisty N. (2016). Analysis of chromosomal aberrations and recombination by allelic bias in RNA-Seq. Nature Communications, 7, 12144.

+ Open protocol

+ Expand

Metformin and FTI Effects on HGPS MSCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

The MSCs derived from HGPS iPS cells (HGPS MSCs) and WT iPS cells (WT MSCs) were cultured in KnockOut Dulbecco’s modified Eagle’s medium (Invitrogen, Carlsbad, CA, USA), supplemented with 20% fetal bovine serum, research grade (Sigma), 1% nonessential amino acids (Invitrogen), 1% glutamax (Invitrogen) and 0.1% β-mercaptoethanol (Invitrogen). Six hours after seeding, the MSCs were treated with 0.1% dimethyl sulfoxide, 1 μmol/l FTI (tipifarnib, R115777; Selleck Chemicals, Houston, TX, USA) or different concentrations of metformin (1,1-dimethylbiguanide hydrochloride, Sigma). metformin treatment was repeated 24 h after the initial treatment. The cells were analyzed after 48 h of treatment.

Egesipe A.L., Blondel S., Lo Cicero A., Jaskowiak A.L., Navarro C., Sandre-Giovannoli A.D., Levy N., Peschanski M, & Nissan X. (2016). Metformin decreases progerin expression and alleviates pathological defects of Hutchinson–Gilford progeria syndrome cells. NPJ Aging and Mechanisms of Disease, 2, 16026-.

+ Open protocol

+ Expand

Feeder-free Culture of Mouse ESCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

E14Tg2a mouse ESCs, a kind gift from Youn, H. D (Seoul National University), were cultured on 0.1% gelatin-coated dishes under feeder-free conditions in KnockOut Dulbecco’s modified Eagle’s medium (Gibco/Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 15% fetal bovine serum (Atlas Biologicals, Fort Collins, CO, USA), 2 mM L-glutamine, 100 U/mL penicillin, 100 µg/mL streptomycin, 100 µM 2-mercaptoethanol, 20 µg/mL ciprofloxacin, 1× non-essential amino acids, and 1000 U/mL leukemia inhibitory factor (LIF; MTI-GlobalStem, Gaithersburg, MD, USA). Stable E14Tg2a mouse ESCs with knockdown or overexpression of Brpf3 were cultured in medium supplemented with 2 µg/mL puromycin. 293T cells (ATCC) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum, 100 U/mL penicillin, and 100 µg/mL streptomycin. All cell lines were tested for mycoplasma contamination.

Cho H.I., Kim M.S., Lee J., Yoo B.C., Kim K.H., Choe K.M, & Jang Y.K. (2020). BRPF3-HUWE1-mediated regulation of MYST2 is required for differentiation and cell-cycle progression in embryonic stem cells. Cell Death and Differentiation, 27(12), 3273-3288.

+ Open protocol

+ Expand

Generation of Tet1/2/3 Triple Knockout mESCs

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

mESCs (E14) were cultured on MEFs in Knock-out Dulbecco’s Modified Eagle’s Medium (Gibco), supplemented with 15% fetal bovine serum (Omega), 0.5% penicillin-streptomycin (Gibco), 0.1 mM non-essential amino acids (Gibco), 0.1 mM 2-mercaptoetanol (Sigma), and 103 U/mL of leukemia inhibitory factor (Millipore). Tet1/2/3 triple knock-out mESCs were generated by the CRISPR/Cas9 technology as described previously^{44 (link)} with slight modifications. Tet1/2/3 sgRNAs were cloned into PX458 (Addgene 48138). Three sgRNAs were transfected simultaneously into mESCs using the iMfectin DNA transfection reagent (Gendepot). mESCs transfected with the vector PX458 without sgRNA were used as control. GFP positive cells were sorted into 96-well plates by flow cytometry and individual colonies were genotyped after 7-day culture.

Fang S., Li J., Xiao Y., Lee M., Guo L., Han W., Li T., Hill M.C., Hong T., Mo W., Xu R., Zhang P., Wang F., Chang J., Zhou Y., Sun D., Martin J.F, & Huang Y. (2019). Tet inactivation disrupts YY1 binding and long-range chromatin interactions during embryonic heart development. Nature Communications, 10, 4297.

+ Open protocol

+ Expand

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?

Revolutionizing how scientists
search and build protocols!