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Sample GSM2474142 Query DataSets for GSM2474142
Status Public on Feb 02, 2017
Title SCC_H3K27me3_ChIPSeq
Sample type SRA
 
Source name Lung tumors
Organism Mus musculus
Characteristics tissue: Lung tumors
tumor subtype: Squamous cell carcinoma
background: FSF:KrasG12D/+; FSF:R26:CreERT2; Lkb1flox/flox
antibody: H3K27me3 (CST C36B11)
Extracted molecule genomic DNA
Extraction protocol Lung tumors were pulverized, cross-linked with 1% formaldehyde in PBS for 10 min at RT, washed in 5mg/ml BSA in PBS and then in just cold PBS, re-suspended in lysis buffer [50mM Tris-HCl, pH 8.1, 10mM EDTA, 1% SDS, 1X complete protease inhibitors (Roche)], and sonicated with the Covaris E210 or S2 sonicator to obtain chromatin fragment lengths of 200-to-1500bp judged by Bioanalyzer DNA High sensitivity kit (Agilent). Fragmented chromatin was diluted in IP buffer [20mM Tris-HCl pH 8.1, 150mM NaCl, 2mM EDTA, 1% Triton X-100] and incubated overnight at 4ºC with Protein G magnetic beads (Dynabeads: Life Technologies) that had been pre-incubated with antibodies. Immunoprecipitates were washed six times with wash buffer [50mM HEPES pH 7.6, 0.5M LiCl, 1mM EDTA, 0.7% Na deoxycholate, 1% NP-40] and twice with TE buffer. Immunoprecipitated (or no IP input) DNA was treated with RNase A and Proteinase K on the beads, recovered in 1% SDS and 0.1M NaHCO3 over a period of 5 hours at 65ºC, column purified with DNA clean and concentrator -25 (Zymo Research).
After a sonication to enrich DNA fragment lengths between 100-300bp, 5 to 10ng of DNA were used for the library construction using NEBNext Ultra DNA Library Prep Kit (NEB E7370). Sequencing was performed on a NextSeq (Illumina) for 38 nucleotides from paired ends according to manufacturer’s instructions.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina NextSeq 500
 
Description st_hw_2893-Sq_1_K27me3
Data processing ChIP-seq paired reads were aligned to the mm10 genome assembly using Bowtie2 with -k 1 option
H3K4me3 and H3K27Ac peaks were called using MACS (v1.4.2) at p-value threshould of 1E-05 with no-ChIP input sample as control.
Wiggle files with a 10bp resolution for H3K27me3 were generated by MACS v1.4.2 with tag shift and then rescaled to a larger of total number of uniquely alignable sequences in two datasets.
Genome_build: mm10
 
Submission date Feb 01, 2017
Last update date May 15, 2019
Contact name Hideo Watanabe
E-mail(s) hideo.watanabe@mssm.edu
Organization name Icahn School of Medicine at Mount Sinai
Department Department of Medicine, Division of Pulmonary, Critical Care and Sleep Medicine
Lab Hideo Watanabe
Street address One Gustave L. Levy Place
City New York
State/province NY
ZIP/Postal code 10029
Country USA
 
Platform ID GPL19057
Series (1)
GSE94365 Lkb1 inactivation drives lung cancer lineage switching governed by Polycomb Repressive Complex 2
Relations
BioSample SAMN06287504
SRA SRX2531392

Supplementary file Size Download File type/resource
GSM2474142_st_hw_2893sq_1_k27me3-noinput-p6-swig_treat_afterfiting_all.rescaled.wig.gz 118.4 Mb (ftp)(http) WIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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