Random differentiation of mouse embryonic stem cell
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE3527 | GEO | 2005/11/01
SECONDARY ACCESSION(S): PRJNA104823
REPOSITORIES: GEO
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Project description:This experiment is part of the FunGenES project (FunGenES - Functional Genomics in Embryonic Stem Cells partially funded by the 6th Framework Programme of the European Union, http://www.fungenes.org). The experiment was conducted at Aventis Pharma, Paris, France. Goal of the experiment is the characterisation of gene expression profile during neuronal differentiation of Sox1Tv2 ES cells in the embryoid body protocol. Materials and methods:Sox1Tv2-E14 derived ES cells have been cultured without serum in liquid culture in order to form embryoid bodies. A 48 hour retinoic acid stimulation has been applied between days 4 and 6 (EB6) for half the samples. After 8 days of differentiation (EB8), EBs have been dissociated and cells plated on poly-D-lysine/laminin coated dishes and cultured in differentiation medium. bFGF has been added to the culture medium during the first 2 days after plating (N2). After removal of bFGF, cells were allowed to differentiate for 4 more days (N6). Protocol described in FunGenES handbook. Relationships between samples: Different time points during differentiation process: ES, EB3, EB4, EB6 (ctl and RA), EB8 (ctl and RA), N2 (ctl and RA), N6 (ctl and RA),Treatments: Retinoic acid treatment: EB4 to EB6bFGF addition (10ng/ml final): postplating to N2. Culture conditions: Liquid medium culture: GMEM + KOSR Differentiation medium: DMEM/F12 and NBM medium with N2 and B27 supplements.
2009-07-10 | E-TABM-669 | biostudies-arrayexpress
Project description:This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/We established protocols to generate monocytes and macrophages from murine embryonic stem cell (129 background) in vitro. Briefly, murine embryonic stem cells were grown in bacteriological dishes in the presence of differentiation medium supplement ed with 15% L929 and 1ng/ml IL-3 for 8 days to generate embryoid bodies (EBs). Subsequently, EBs were transferred to gelatinized tissue culture dish. Supernatants containing macrophage progenitors were collected, spun down and plated onto bacteriological dishes to differentiate into macrophages. Four biological replicates of murine embryonic stem cells, macrophage progenitors and macrophages were collected and their RNA were isolated via the Qiagen RNA Isolation Kit. The RNA were treated with RNAse free DNase and eluted in RNase-free water. THe RNA concentration was assessed using a NanoDrop spectrophotometer, while RNA integrity (RIN >8) were obtained for all samples by Agilent 2100 Bioanalyzer. We aim to sequence 5GBp per replicate and 4 replicates per lane on an Illumina platform.
2013-05-28 | E-ERAD-132 | biostudies-arrayexpress
Project description:time-course experiment with embryoid bodies of CGR8 mouse embryonic stem cells ; in the whole time-series RNA from 0 days old embryoid bodies were hybridized against RNA from 3 days and 10 days old embryoid bodies Keywords = embryoid bodies Keywords = mouse Keywords = time-course Keywords = oligonucleotide array Keywords: time-course
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Project description:iPSCs were maintained on Geltrex coated flat culture dish in E8 media according to manufacturer’s guidelines. Colonies were manually harvested at 60-80% confluence. Cells were then collected and dissociated into single cells using EDTA. Cells were put on to ultra low attachment 24 well or 96 well plate to allow them to form aggregated in suspension with ROCK inhibitor. Cell aggregates 191were cultured in E8 medium with daily medium change for 6-7 days. Control iPSC-A (iPSC-aggregates) were plated on a Geltrex in 96 well plate or 8 well culture chamber. And then aggregates were treated E8 medium with daily medium change for 12-14 days.
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Project description:Synovial fibroblasts were extracted from knee joints of naïve mice(DBA/1J, male 8-12 week old), and passaged 3-4 times. Cells were plated at a concentration of 1 X 106 cells/dish in 60 mm dish in 5 ml of RPMI1640 medium with 1% heat-inactivated FCS at 37°C, 5% CO2. Cells were treated vehicle or stimulated by IL-1beta. In IL-1beta(5ng/ml) stimulated groups, indomethacin (1 μM) and iloprost (1μM) were further added either alone or in combination. After 6 hour stimulation as described above, cells were isolated and frozen by liquid nitrogen, and stored at –80°C before use. Total RNA was extracted by RNAeasy Mini Kit. The RNA sample was labeled for hybridization onto the genechip array according to the standard affymetrix protcols. Keywords = arthritis prostacyclin Keywords: other
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Project description:GBM cell lines were plated onto 10 cm dishes at 100,000-200,000 cells per dish and allowed to grow for 3-4 days until confluency reached 50-70%. Media for all cell lines was DMEM with 10% FBS including 25 mM glucose, 6.2 mM glutamine and 200 uM Oleic Acid (conjugated to BSA). 1-2 hours prior to tracer incubation, media was aspirated and fresh media was used. Immediately prior to tracer incubation, media was again aspirated and then cells were washed with warm PBS to remove unlabeled metabolites. Cells were then incubated with labeled tracer (DMEM with 10% FBS including 25 mM Uniformly labeled 13C-6 glucose, 6.2 mM unlabeled glutamine and 200 uM unlabeled oleic acid). After 2 hours of incubation with tracer, media was aspirated, cells were quickly washed with 15 mL DI water, quenched with liquid nitrogen and then stored at -80oC until analysis.
2015-03-17 | ST000301 | MetabolomicsWorkbench
Project description:To unravel the molecular mechanism by which HOXB4 promotes the expansion of early hematopoietic progenitors within differentiating ES cells, we analzed the gene expression profiles of embryoid bodies (EBs) in which transcription of HOXB4 had been induced or not induced. A substantial number of the identified HOXB4 target genes are involved in signaling pathways important for controlling self-renewal, maintenance and differentiation of stem cells. Furthermore, we demonstrate that HOXB4 activity and FGF-signaling are intertwined. HOXB4-mediated expansion of ES cell-derived early progenitors was enhanced by specific and complete inhibition of FGF-receptors. In contrast, the expanding activity of HOXB4 on hematopoietic progenitors in day4-6 embryoid bodies was blunted in the presence of basic FGF (FGF2) indicating a dominant negative effect of FGF-signaling on the earliest hematopoietic cells. Taken together, we show that modulation of FGF signaling is an essential feature of HOXB4 activity in the context of embryonic hematopoiesis. Experiment Overall Design: The Hoxb4i ES cell line (Kyba et al. 2002, Cell 109:29-37) contains an integrated âtet-onâ cassette that allows induction of HOXB4 expression upon treatment with doxycycline. These ES cells can be used to produce hematopoietic cells through the formation of embryoid bodies (EBs). Hematopoiesis starts in these EBs at day 4 and the differentiation into hematopoietic fates can be quantified by colony assays on methyl-cellulose using cells dissociated from EBs at day 6 of incubation. The induction of HOXB4 by incubation with doxycycline increases the production of hematopoietic progenitors within EBs by day 6. Using this specific ES cell line, we compared the transcriptome between embryoid bodies (EBs) in which transcription of HOXB4 had been induced or not induced from day 4 to day 6 (48hours). Experiment Overall Design: Biological replicates: 3
2008-06-16 | E-GEOD-9044 | biostudies-arrayexpress
Project description:We used high throughput miRNA sequencing to investigate differences between two mESC lines (D3 and B8 ESCs). Undifferentiated ESCs, cells differentiating in embryoid bodies (day 7), and also cells from embryoid bodies outgrowth (day 21) were analyzed.
2018-01-18 | GSE102719 | GEO
Project description:C2C12 mouse myoblasts (American Type Culture Collection, Manassas, VA) were maintained in growth medium (DMEM supplemented with 10% Fetal Bovine serum, Invitrogen Corporation, Carlsbad, CA) in 5% CO2. Cells were plated at a density of 40-50% in 100mm dishes. When cells reached ~90% confluency 24 hours later, they were switched to differentiation media (DMEM containing 2% horse serum; GIBCO, Invitrogen Corporation, Carlsbad, CA) which was subsequently changed every 48 hours. All cells were allowed to differentiate for 4 days with feeding every two days (day 0, 2, and 4). Beginning at day 4, control cells were re-fed every 48 hours whereas experimental cells were not re-fed. After an additional 4 days (8 days post differentiation), cells were harvested and total RNA was isolated from control and "starved" (non re-fed cells) Keywords: repeat sample
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