NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM4105815 Query DataSets for GSM4105815
Status Public on Oct 03, 2019
Title MCF7_shARID1A_EZH1
Sample type genomic
 
Channel 1
Source name EZH1 ChIP DNA from MCF7_shARID1A
Organism Homo sapiens
Characteristics cell type: Breast cancer cell line
Extracted molecule genomic DNA
Extraction protocol 1×10E7 cells were cross-linked in fixing solution. After 15 min incubation at room temperature, cross-linking was stopped by adding 1.375 M glycine. The cells were centrifuged for 5 min, 1,400 × g at 4°C. The supernatant was discarded and the pellet was washed two times in PBS. After the washes, cell pellets were resuspended in LB1 (50 mM Hepes-KOH (pH 7.5), 140 mM NaCl, 1 mM EDTA, 10% Glycerol, 0.5% IGEPAL CA-630, 0.25% Triton X-100) and incubated for 10 min on ice. After centrifugation for 5 min, 1,400 × g at 4°C, the pellet was resuspended in LB2 (10 mM Tris-HCl (pH 8.0), 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA) for 10 min at room temperature. After centrifugation for 5 min, 1,400 × g at 4°C, the nuclei were resuspended in micrococcal nuclease digestion buffer and incubated in 37°C to digest chromatin to the appropriate size fragments. Digestion was stopped by adding 0.5 M EDTA. After centrifugation, nuclei pellets were resuspended in LB3 (10 mM Tris-HCl (pH 8.0), 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Sodium Deoxycholate, 0.5% N-Lauroylsarcosine) and sonicated for 10 min. After sonication and 10 min high-speed centrifugation, sheared chromatin was recovered in the supernatant.
Label Cy5
Label protocol Labeled DNA was prepared from 2.6 μg of amplified DNA using SureTag DNA Labeling Kit according to the manufacturer's instructions.
 
Channel 2
Source name Input DNA from MCF7_shARID1A
Organism Homo sapiens
Characteristics cell type: Breast cancer cell line
Extracted molecule genomic DNA
Extraction protocol 1×10E7 cells were cross-linked in fixing solution. After 15 min incubation at room temperature, cross-linking was stopped by adding 1.375 M glycine. The cells were centrifuged for 5 min, 1,400 × g at 4°C. The supernatant was discarded and the pellet was washed two times in PBS. After the washes, cell pellets were resuspended in LB1 (50 mM Hepes-KOH (pH 7.5), 140 mM NaCl, 1 mM EDTA, 10% Glycerol, 0.5% IGEPAL CA-630, 0.25% Triton X-100) and incubated for 10 min on ice. After centrifugation for 5 min, 1,400 × g at 4°C, the pellet was resuspended in LB2 (10 mM Tris-HCl (pH 8.0), 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA) for 10 min at room temperature. After centrifugation for 5 min, 1,400 × g at 4°C, the nuclei were resuspended in micrococcal nuclease digestion buffer and incubated in 37°C to digest chromatin to the appropriate size fragments. Digestion was stopped by adding 0.5 M EDTA. After centrifugation, nuclei pellets were resuspended in LB3 (10 mM Tris-HCl (pH 8.0), 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% Sodium Deoxycholate, 0.5% N-Lauroylsarcosine) and sonicated for 10 min. After sonication and 10 min high-speed centrifugation, sheared chromatin was recovered in the supernatant.
Label Cy3
Label protocol Labeled DNA was prepared from 2.6 μg of amplified DNA using SureTag DNA Labeling Kit according to the manufacturer's instructions.
 
 
Hybridization protocol Hybridization was performed at 65°C for 40h in a rotating Agilent hybridization oven. After hybridization, microarrays were washed at room temperature with Agilent Oligo aCGH/ChIP-on-chip Wash Buffer 1 for 5min and with Agilent Oligo aCGH/ChIP-on-chip Wash Buffer 2 for 5min at 37°C.
Scan protocol Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner C using the two color scan setting for 2x400K microarray slides (Scan Area: 61x21.6 mm, Scan resolution: 3 um, Dye channel: red and green, Tiff: 16bit, XDR ratio: NoXDR)
Data processing The scanned images were analyzed with Feature Extraction Software 10.7 using default parameters (protocol: ChIP_107_Sep09) to obtain background subtracted and spatially detrended Processed Signal intensities.
 
Submission date Oct 02, 2019
Last update date Oct 03, 2019
Contact name Makoto Yamagishi
E-mail(s) [email protected]
Organization name The University of Tokyo
Lab Lab. Tumor Cell Biology
Street address 4-6-1, Shirokanedai
City Minato-ku
State/province Tokyo
ZIP/Postal code 108-8639
Country Japan
 
Platform ID GPL19784
Series (1)
GSE138342 Epigenetic pattern after EZH1,2 inhibition in lymohoma cells

Supplementary file Size Download File type/resource
GSM4105815_MCF7_shARID1A_EZH1.txt.gz 43.2 Mb (ftp)(http) TXT
Processed data are available on Series record

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