Comment[ArrayExpressAccession]	E-GEOD-63304
MAGE-TAB Version	1.1		
Public Release Date	2014-11-26
Investigation Title	Epigenetic inheritance uncoupled from sequence-specific recruitment		
Comment[Submitted Name]	Epigenetic inheritance uncoupled from sequence-specific recruitment
Experiment Description	We used ChIP-seq to examine the requirements for inheritance of H3K9me at endogenous and ectopic loci. H3K9me2 and H3K9me3 ChIP-seq in multiple S. pombe strains		
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	Jih	Jih	Moazed
Person First Name	Gloria	Gloria	Danesh
Person Mid Initials		T	
Person Email	gjih@fas.harvard.edu		
Person Affiliation	Harvard Medical School		
Person Address	Cell Biology, Harvard Medical School, 240 Longwood Ave, LHRRB 517, Boston, MA, USA		
Person Roles	submitter		
Protocol Name	P-GSE63304-1		
Protocol Description	Fixed cells were resuspended in 500 ul lysis buffer (50 mM Hepes-KOH, pH 7.5, 500 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, and protease inhibitors) and disrupted by bead beating (Mini Beadbeater, Biospec products) for 6x30 sec with 1 ml 0.5 mm glass beads. Tubes were punctured and the flow-through was collected in a new tube by centrifugation. After sonication for 3x20 sec at 40% amplitude (Branson Digital Sonifier), the extract was centrifuged (Eppendorf 5415R) for 15 min at 13000 rpm. Antibody was pre-incubated with washed Dynabeads Protein A, and for each immunoprecipitation, 2 ug antibody coupled to 30ul beadsM- was added to 400 ul soluble chromatin, and lysis buffer was added for a final volume of 600 ul. Samples were rotated for 2h at 4M-0C, the beads were collected on magnetic stands, and washed 3 times with 1 ml lysis buffer and once with 1 ml TE, and eluted with 100 ul pre-heated buffer (50 mM Tris pH 8.0, 10mM EDTA, 1% SDS) at 65M-0C for 15 min. The eluate was collected and 150 ul 1xTE/0.67% SDS was added. Input and immunoprecipitated samples were incubated overnight at 65M-0C to reverse crosslinks, and treated with 50 ug RNase A at 37M-0C for 1h. DNA was cleaned up with QIAquick PCR Purification Kit (Qiagen). Libraries for Illumina sequencing were constructed following the manufacturerM-bM-^@M-^Ys protocols, starting withM- M-bM-^HM-<5 ng of immune-precipitated DNA fragments. Each library was generated with custom-made adapters carrying unique barcode sequence at the ligating end (Wong and Struhl, 2011). Barcoded libraries were mixed and sequenced with Illumina HiSeq2000. Barcodes are 6/8/9 nt in length and should be followed by a T.		
Protocol Type	nucleic acid library construction protocol		
Experimental Factor Name	strain	media	antibody
Experimental Factor Type	strain	media	antibody
Comment[SecondaryAccession]	GSE63304		
Comment[GEOReleaseDate]	2014-11-26		
Comment[ArrayExpressSubmissionDate]	2014-11-14		
Comment[GEOLastUpdateDate]	2014-11-26		
Comment[AEExperimentType]	ChIP-seq		
Comment[SecondaryAccession]	SRP049816		
SDRF File	E-GEOD-63304.sdrf.txt