NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM486274 Query DataSets for GSM486274
Status Public on Mar 22, 2010
Title GM18505_NFkB_II_ChIPSeq_Rep1
Sample type SRA
 
Source name Lymphoblastoid Cells
Organism Homo sapiens
Characteristics cell line: GM18505
cell type: Lymphoblastoid cell
treatment: TNF-alpha induced (6h)
chip antibody: NFκB p65 (C-20) rabbit polyclonal antibody
Biomaterial provider Coriell Cell Repositories http://ccr.coriell.org/Sections/Search/Search.aspx?PgId=165&q=GM18505
Treatment protocol Prior to cross-linking, cells were treated with 25 ng/mL human recombinant TNF-alpha (eBioscience #14-8329) for six hours at 37°C, 5% CO2.
Growth protocol Lymphoblastoid cell lines were grown in RPMI-1640 Medium, supplemented with glutamine, 15% FBS, and antibiotics (penicillin-streptomycin) at 37C in 5% CO2, to a density of 6-8 x10^5 cells/ml.
Extracted molecule genomic DNA
Extraction protocol Chomatin immunoprecipitation was performed as previously described (Euskirchen et al. 2007; Rozowsky et al. 2009). 2 x 10^8 cells were cross-linked in 1% formaldehyde for 10 minutes at room temperature. The cross-linking reaction was quenched by adding glycine to a final concentration of 125 mM. Nuclear lysates were sonicated using a Branson 250 Sonifier (power setting 2, 100% duty cycle for 7 × 30-s intervals), such that the chromatin fragments ranged from 50-2000kb. Clarified lysates were divided in half and treated overnight at four degrees Celsius with 8μg of either anti-NFκB p65 (C-20) rabbit polyclonal antibody (Santa Cruz sc-372) or normal rabbit IgG (Santa Cruz Sc-2027). Protein-DNA complexes were captured on Protein A agarose beads (Upstate #16-156) and eluted in 1% SDS TE buffer at 65°C. Following cross-link reversal and purification, the ChIP DNA sequencing libraries were generated according to Illumina DNA Sample Kit Instructions (Illumina Part # 0801– 0303). The protocol was modified such that enzymes were obtained from other suppliers, as described in Auerbach et al. 2009.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina Genome Analyzer II
 
Description Chromatin IP against NFκB p65 (C-20) rabbit polyclonal antibody
Data processing FASTQ files are either the sequence.txt files from the Illumnia pipeline (which exclude low quality reads) or, when available, are generated from the Illumina export.txt files which include all reads. Both types of alignment files, eland_result.txt and eland_multi.txt, are directly out of the Illumnia pipeline and unmodified. The narrowPeak files are the scored results generated from the PeakSeq pipeline (http://www.ncbi.nlm.nih.gov/pubmed/19122651?dopt=Abstract) and formatted in the UCSC narrowPeak format.
 
Submission date Dec 15, 2009
Last update date May 15, 2019
Contact name Flora Vaccarino
Organization name Yale University
Department Child Study Center
Lab Vaccarino
Street address 230 South Frontage Road
City New Haven
State/province CT
ZIP/Postal code 06520
Country USA
 
Platform ID GPL9115
Series (2)
GSE19485 Human variation in PolII and NF-KappaB binding (ChIP-seq study with NF-KappaB)
GSE19486 Human variation in PolII and NF-KappaB binding
Relations
SRA SRX017863
BioSample SAMN00010238

Supplementary file Size Download File type/resource
GSM486274_GM18505_IgGwithTNF-NFKB-0626091831_rep1_FC30WBPHM_20090202_s_3_eland_result.txt.gz 279.8 Mb (ftp)(http) TXT
GSM486274_GM18505_IgGwithTNF-NFKB-0626091831_rep1_narrowPeak.bed.gz 68.8 Kb (ftp)(http) BED
SRA Run SelectorHelp
Processed data provided as supplementary file
Raw data are available in SRA

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap