characteristics.text: 2013-02-08T17:03:00Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T16:09:43Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA1712777
characteristics.characteristic: External Id
characteristics.text: Pediatric Diabetes Research Center, UCSD Stem Cell Program
characteristics.characteristic: INSDC center name
characteristics.text: 2013-02-08T17:03:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T16:09:43Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS197531
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-1086:DP1
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: Cyt49
characteristics.characteristic: cell line
characteristics.text: embryonic stem cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0002322
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: CyT49 hESCs were maintained and differentiated as previously described [DAmour, K.A., et al., Nat Biotechnol, 2006. 24:1392-401; Kroon, E., et al., Nat Biotechnol, 2008. 26:443-52; Kelly, O.G., et al., Nat Biotechnol. 29:750-6]. Non-adherent conditions were applied to hESC differentiation. Chromatin immunoprecipitations were performed on aggregated hESC-derived cells, FACS-sorted cells and transplanted grafts as previously described [Bhandare, R., et al., Genome Res. 20:428-33]. Briefly, samples were crosslinked in 1.1% formaldehyde/PBS for 10 min at room temperature and then quenched with 0.125M glycine/PBS. Samples were subsequently washed twice with PBS and then lysed in 1% SDS. For sonication, lysates were sonicated with a Bioruptor Sonicator (Diagenode) three times, for 5 min each with a 30 sec on and 30 sec off cycle, resulting in 200-500bp chromatin fragments. The sheared chromatin was incubated overnight at 4 C with 50 ul of Dynabeads sheep anti-rabbit IgG (Invitrogen), preincubated with 5 ug rabbit anti-H3K4me3 (Millipore, 04-745) or rabbit anti-H3K27me3 (Millipore, 07-499) antibodies. Immnuoprecipitated complexes were further eluted, reverse crosslinked, and subjected to library preparation. For aggregated hESC-derived and FACS-sorted samples, a total of 2 to 5 _ 106 cells were used for each ChIP-seq run. All ChIP-seq libraries were prepared as per Illuminas instructions (http://www.illumina.com). Briefly, total ChIP DNA was subjected to end-repair and ligated to the Adaptor Oligo Mix (Illumina). For input library preparation, 10 ng of input DNA from each sample was used. After adaptor ligation, DNA fragments were size-fractionated by gel electrophoresis and excised at (+25 bp,-25bp). Following gel purification, DNA fragments were amplified with 18 PCR cycles and purified using a MiniElute PCR Purification kit (Qiagen).
characteristics.characteristic: description
characteristics.text: embryo
characteristics.characteristic: developmental stage
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: DP1
characteristics.characteristic: title