characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2015-07-24T17:01:24Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T21:49:23Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA3157841
characteristics.characteristic: External Id
characteristics.text: Pfizer Neusentis
characteristics.characteristic: INSDC center name
characteristics.text: 2015-07-24T17:01:24Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T21:49:23Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS618063
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-3137:IgG
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: epithelial cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0000066
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: RPE were generated from the hESC cell line SHEF1.3 or iPSC reprogrammed from erythroblasts using the spontaneous differentiation method described previously (Vugler et al., 2008). Pluripotent cells were cultured as feeder-free colonies on hESC-qualified Matrigel (BD) in mTesR1 (StemCell Technologies) media. RPE foci were excised with a scalpel and dissociated into a single cell suspension using Accutase (Gibco) and plated onto CellStart (Invitrogen) coated surfaces. ChIP was performed essentially as described before (Sanders et al., 2013). Briefly, 1 million cells were crosslinked with 1% Formaldehyde for 10 minutes and the reaction was quenched with 125mM glycine. Sonication was performed on isolated nuclei using the Bioruptor Plus with cooling system for 35 cycles of each 30 seconds on and 30 seconds off at the maximum setting leading to a fragment size between 200-300bp. For each IP, 50 ul of Protein A Dynabeads were incubated with 5 ug of the appropriate antibody (anti-FOXM1 [Santa Cruz Biotechnology, SC-502] or Rabbit IgG REF) overnight at 4oC. Chromatin immunoprecipitation was performed by adding the 100 ul of preblocked antibody-bead complexes per sample and incubating overnight at 4oC on a rotator. Elution and crosslink reversal of both sample and input was performed by incubating the samples for 6 hours at 65oC in a waterbath. The DNA was purified using the Zymo ChIP DNA Clean & Concentrator kit (Zymo Research) according to the manufacturer's instructions. The sequencing library was generated using the MicroPlex Library Preparation kit (Diagenode) and size selected using Agencourt AMPure XP. The libraries were quantified using the qPCR based Illumina library quantification kit (Kappa Biosystems) .
characteristics.characteristic: description
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: IgG
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005030
characteristics.characteristic: title