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Sample GSM461100 Query DataSets for GSM461100
Status Public on Oct 09, 2009
Title U87-RNApolII ChIP-chip
Sample type genomic
 
Channel 1
Source name U87 RNA polII ChIP Normoxia
Organism Homo sapiens
Characteristics antibody: RNA polymerase II mAb (Abcam, ab5408)
cell line: U87 cells
stress: normoxic (ambient) conditions
Treatment protocol U87 cells were cultured for 4 hours under normoxic or hypoxic (0.5%O2) conditions.
Extracted molecule genomic DNA
Extraction protocol Cells were fixed with 1% formaldehyde (37°C, 10 min) and lysed with 0.5% SDS lysis buffer. Chromatin was then sonicated to 500-1000bp fragments and immunoprecipitation carried out with HIF-1α pAb (Novus Biologicals, NB 100-134). RNA polymerase II and H3K4me3 ChIP were carried out using normoxic U87 or HepG2 cell samples with RNA polymerase II mAb (Abcam, ab5408) and H3K4me3 pAb (Abcam, ab8580). A small aliquot for each biological sample are reserved for an input control. Both Chiped DNA and Input samples were amplified by LM-PCR.
Label biotin
Label protocol Amplified ChIP and Input samples were fragmented with DNase to 50~100bp size and labeled using the Terminal transferase (TdT, Promega) to add biotinylated-ddATP to the ends of the fragments
 
Channel 2
Source name U87 RNA polII input Normoxia
Organism Homo sapiens
Characteristics cell line: U87 cells
stress: normoxic (ambient) conditions
antibody: none
Treatment protocol U87 cells were cultured for 4 hours under normoxic or hypoxic (0.5%O2) conditions.
Extracted molecule genomic DNA
Extraction protocol Cells were fixed with 1% formaldehyde (37°C, 10 min) and lysed with 0.5% SDS lysis buffer. Chromatin was then sonicated to 500-1000bp fragments and immunoprecipitation carried out with HIF-1α pAb (Novus Biologicals, NB 100-134). RNA polymerase II and H3K4me3 ChIP were carried out using normoxic U87 or HepG2 cell samples with RNA polymerase II mAb (Abcam, ab5408) and H3K4me3 pAb (Abcam, ab8580). A small aliquot for each biological sample are reserved for an input control. Both Chiped DNA and Input samples were amplified by LM-PCR.
Label biotin
Label protocol Amplified ChIP and Input samples were fragmented with DNase to 50~100bp size and labeled using the Terminal transferase (TdT, Promega) to add biotinylated-ddATP to the ends of the fragments
 
 
Hybridization protocol Approximately 3ug of DNA was hybridzed per array.
Scan protocol Both hybridization and scan were performed by Microarray Core at Dana-Farber Cancer Institute.
Description U87 RNA polII ChIP Normoxia Biological Rep 1-3 v.s. Inputs, Affymetrix Human Promoter 1.0R Array
test CEL file: U87_PolII_C1.CEL
test CEL file: U87_PolII_C2.CEL
test CEL file: U87_PolII_C3.CEL
control CEL file: U87-NI1.CEL
control CEL file: U87-NI2.CEL
control CEL file: U87-NI3.CEL
Data processing The Model-based Analysis of Tiling-array (MAT) algorithm was used to identify regions enriched by ChIP-chip (ChIP hits). For the U87 HIF-1 ChIP, the triplicate hypoxic U87 HIF-1 ChIP samples were compared directly to triplicate normoxic samples; MAT was run with parameters of bandwidth = 200, maximum gap = 400, minimum probes = 10, and p-value cutoff = 1x10-5. For H3K4me3, H3K27me3, and RNA Pol II ChIP, normoxic ChIP samples were compared to matched input samples; the MAT parameters were increased to account for broader peaks: bandwidth = 500, maximum gap = 400, minimum probes = 20, and p-value cutoff = 1x10-5. The MAT library and mapping files were based on the March 2006 Human Genome Assembly (HG18). Hits flagged by MAT as mapping to repeat regions were excluded from consideration in all cases.
 
Submission date Oct 09, 2009
Last update date Oct 09, 2009
Contact name Xiaobo Xia
E-mail(s) Xiaobo_Xia@dfci.harvard.edu
Organization name Dana-Farber Cancer Institute
Department Pediatric Oncology
Lab Dr. Andrew Kung
Street address 44 Binney St, Mayer 649
City Boston
State/province MA
ZIP/Postal code 02115
Country USA
 
Platform ID GPL5082
Series (2)
GSE18499 Genome-wide HIF-1, RNA polII, and H3K4me3 binding sites in U87 and HepG2 cells
GSE18505 HepG2 hepatoma and U87 glioma cells: transcriptomic and genomic analyses

Supplementary file Size Download File type/resource
GSM461100_U87-NI1.CEL.gz 19.2 Mb (ftp)(http) CEL
GSM461100_U87-NI2.CEL.gz 18.8 Mb (ftp)(http) CEL
GSM461100_U87-NI3.CEL.gz 19.4 Mb (ftp)(http) CEL
GSM461100_U87_PolII_C1.CEL.gz 19.5 Mb (ftp)(http) CEL
GSM461100_U87_PolII_C2.CEL.gz 20.0 Mb (ftp)(http) CEL
GSM461100_U87_PolII_C3.CEL.gz 20.0 Mb (ftp)(http) CEL
GSM461100_U87_polII.Hs_PromPR_v01-3_NCBIv36.NR.bpmap_matscore.bar.gz 20.6 Mb (ftp)(http) BAR
GSM461100_U87_polII.bed.gz 119.4 Kb (ftp)(http) BED
Processed data included within Sample table
Processed data provided as supplementary file

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