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Sample GSM1040719 Query DataSets for GSM1040719
Status Public on Dec 20, 2012
Title Day3_SSEA1+ M2 K4
Sample type SRA
 
Source name Sorted cells
Organism Mus musculus
Characteristics mark: K4
chip antibody: Anti-Trimethyl Histone H3 (Lys4) Antibody, (Millipore, clone: CMA304, catalog#: 05-1339, lot#: 2024897)
time after induction[days]: 3
cell stage: Intermediates
thy1: NEG
ssea1: POS
gfp: NEG
Treatment protocol Harvested cells were incubated with antibodies against Thy1.2 (PE conjugated, 53-2.1, eBiosciences) and SSEA-1 (mouse IgM, MC-480, Developmental Hybridoma Bank) for 20 minutes. Cells were washed in PBS and then incubated for 20 minutes with APC conjugated anti mouse IgM, (eBioscience) and Pacific Blue-conjugated streptavidin (Invitrogen). The cells were washed in PBS, resuspended in propidium iodide 5% FBS/PBS solution and passed through a 40mm cell strainer to achieve single cell suspension. Cells were sorted on a FACSAria (BD Biosciences ) and/or analysed in a LSRII (BD Bioscience). For analysis and/or sorting of intermediates, cells were stained with Thy1.2 and SSEA1 antibodies and sorted or analyzed as indicated.
Growth protocol Mouse Embryonic fibroblast (MEFF) cultures were established from E13.5 embryos from a reprogrammable mice strain carrying one copy of the OKSM cassette and Rosa26-M2rtTA allele (het/het) or carrying two copies of the OKSM cassette and Rosa26-M2rtTA allele (ho/ho). Reprogramming was performed in ESC medium (KODMEM with 15% FBS, L-Glutamin, penicillin-streptomycin, non-essential amino acids, b-mercaptoethanol and 1000 U/ml LIF) in the presence of doxycycline.
Extracted molecule genomic DNA
Extraction protocol iPSCs were harvested when they reached about 50% confluency and preplated on non-gelatinized T25 flasks for 45 minutes to remove feeder cells. Intermidiates cell types were harvest after FACS sorting.
ChIP complexes were eluted with 1% SDS in 100 mM NaHCO3 and crosslinks were reversed by incubating samples over night at 65C.
DNA fragments were purified using the QIAquick PCR Purification Kit Cat number: 28104.
Libraries for sequencing were prepared using ChIP-Seq Sample Prep Kit from Illumina, Cat IP-102-1001
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2000
 
Description Sample name: KH181.K4
ChIPed DNA
*p.cov.bigWig: processed data file (plus strand).
*m.cov.bigWig: processed data file (minus strand).
Data processing alignment using bwa
coverage using genomeCoverageBed in bedtools
conversion of coverage to bigWig format using bedGraphToBigWig
Genome_build: mm9
Supplementary_files_format_and_content: content is coverage (i.e., number of reads aligning at each position in the genome) and format is bigWig (see genome.ucsc.edu/goldenPath/help/bigWig.html)
 
Submission date Nov 23, 2012
Last update date May 15, 2019
Contact name Ben S. Wittner
E-mail(s) wittner.ben@mgh.harvard.edu
Organization name Massachusetts General Hospital
Department Center for Cancer Research
Lab Lawrence
Street address 149 13th Street
City Boston
State/province MA
ZIP/Postal code 02129
Country USA
 
Platform ID GPL13112
Series (2)
GSE42477 Defining a molecular roadmap of cellular reprogramming into iPS cells [ChIP-Seq]
GSE42478 Defining a molecular roadmap of cellular reprogramming into iPS cells
Relations
SRA SRX206138
BioSample SAMN01818503
Named Annotation GSM1040719_EAa69.m.cov.bigWig
Named Annotation GSM1040719_EAa69.p.cov.bigWig

Supplementary file Size Download File type/resource
GSM1040719_EAa69.m.cov.bigWig 234.4 Mb (ftp)(http) BIGWIG
GSM1040719_EAa69.p.cov.bigWig 234.4 Mb (ftp)(http) BIGWIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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