Comment[ArrayExpressAccession]	E-GEOD-47483
MAGE-TAB Version	1.1		
Public Release Date	2013-07-22
Investigation Title	H3K27me3 is maintained at a reduced level in Suz12(Bgal/Bgal) ESCs [ChIP-Seq]		
Comment[Submitted Name]	H3K27me3 is maintained at a reduced level in Suz12(Bgal/Bgal) ESCs [ChIP-Seq]
Experiment Description	Suz12(Bgal/Bgal) ESCs express a truncated form of Suz12 fused to Beta-galactosidase. These cells maintain a reduced level of H3K27me3 despite this mutation to a core component of PRC2, unlike Eed-/- ESCs whose H3K27me3 is ablated. Two ESC lines mutant in genes of core components of Polycomb Repressive Complex 2 were assessed for H3K27me3 by ChIP-seq, as compared to a wild type ESC line.		
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl	
Person Last Name	Boyer	Thornton	Boyer
Person First Name	Laurie	Seraphim	Laurie
Person Mid Initials	A	R	A
Person Email	lboyer@mit.edu		
Person Affiliation	Massachusetts Institute of Technology		
Person Phone	617 324-3335		
Person Fax	617 253-8699		
Person Address	Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA, USA		
Person Roles	submitter		
Protocol Name	P-GSE47483-3	P-GSE47483-2	P-GSE47483-1
Protocol Description	Illumina Offline BaseCaller1.9.3 software used for basecalling. ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie 0.12.3, with the following parameters --solexa1.3-quals --best --strata -m 1 -n 2 -p 8 -S. Sequences were extended +200 bp for histone marks and allocated in 25-bp bins. A Poissonian model was used to determine statistically enriched bins with a P-value threshold set at 1x10-9 as described previously (Marson et al., 2008).  Additionally, we required that genomic bins were at least 5 fold over input to be considered enriched peaks.  Genome_build: mm9 Supplementary_files_format_and_content: wig files were generated using an in-house perl script interacting with Bedtools (as described in Wamstad et al. Cell 2012).	The ChIP protocol has been adapted from previous studies (Lee et al., 2006; Boyer et al., 2006, Creyghton et al., 2008) with the following modification: Diagenode Bioruptor was used to sonicate with 30 cycles of 30 sec on (high), 30 sec off. The samples were sonicated in 15ml polystyrene tubes at 4°C while samples were immersed in ice cold water. Millipore 07-449 antibody was used in the ChIP. The ChIP DNA was then used to prepare sequencing libraries as described in Schmidt et al., 2009. For this study, a 1:40 dilution of Single End read adapters (Illumina) was used in the ligation reaction. Each sample was amplified for 18 cycles. The samples were then examined for proper size and structure by Bioanalyzer (Agilent) and qPCR.	ESCs were plated with irradiated murine embryonic fibroblasts (MEFs) and grown under typical ESC conditions on gelatinized tissue culture plates. Briefly, cells were grown in Knockout DMEM supplemented with 15% fetal bovine serum, leukemia inhibitory factor (LIF), non-essential amino acids, L-glutamine, and penicillin/streptomycin as previously described (Boyer et al., 2006). Cells were then preplated - plated on cell culture plates without gelatin for 20 minutes to allow the MEFs to attach and thus deplete the cell population of MEFs - before collection for ChIP.
Protocol Type	normalization data transformation protocol	nucleic acid library construction protocol	growth protocol
Experimental Factor Name	STRAIN OR LINE	GENOTYPE	
Experimental Factor Type	strain or line	genotype	
Comment[SecondaryAccession]	GSE47483		
Comment[GEOReleaseDate]	2013-07-22		
Comment[ArrayExpressSubmissionDate]	2013-05-29		
Comment[GEOLastUpdateDate]	2013-07-22		
Comment[AEExperimentType]	ChIP-seq		
Comment[SecondaryAccession]	SRP023268		
Comment[SequenceDataURI]	http://www.ebi.ac.uk/ena/data/view/SRR869295-SRR869297		
SDRF File	E-GEOD-47483.sdrf.txt