 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Feb 02, 2012 |
| Title |
Human_meth_hg17 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
MeDIP DNA of peripheral blood cells from adult human females
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: peripheral blood cells
antibody: anti-5-methylcytosine antibody
antibody vendor: Aviva System Biology
antibody catalog#: AMM99021
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
A mixture of genomic DNA of peripheral blood cells from four human or four chimpanzee female individuals (a total of 5 μg) was fragmented by sonication into a 0.3- to 1-kb size range, denatured at 95˚C for 10 min, and cooled on ice. The denatured DNA was incubated with 5 μg of anti-5-methylcytosine antibody (Aviva Systems Biology, San Diego, CA) in 200 μl of IP buffer (10 mM sodium-phosphate, 140 mM NaCl, 0.05% Triton-X100 at pH7.0) for 3.5 hours at 4˚C. The antibody-bound DNA fragments were magnetically collected after incubation with Dynabeads M-280 (Life Technologies, Grand Island, NY) coated with anti-mouse IgG antibody for 3 hours at 4˚C. The beads were washed 5 times with 700 μl of IP buffer, resuspended in 250 μl of lysis buffer (50 mM TrisHCl at pH 8.0, 10 mM EDTA, 0.5% SDS) containing Proteinase K. After incubation at 55˚C for 30 min, DNA was recovered by phenol-chloroform extraction and subsequent ethanol precipitation.
|
| Label |
biotin
|
| Label protocol |
The immunoprecipitated DNA was amplified by WGA2 (Sigma Aldrich, Saint Louis, MO) to give a total of ~10 μg DNA, and then labeled with biotine-dUTP using GeneChip WT double stranded DNA terminal labeling kit (Affymetix, Santa Clara, CA).
|
| |
|
| Channel 2 |
| Source name |
Input DNA of peripheral blood cells from adult human females
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: peripheral blood cells
antibody: none, input
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
A mixture of genomic DNA of peripheral blood cells from four human or four chimpanzee female individuals (a total of 5 μg) was fragmented by sonication into a 0.3- to 1-kb size range, denatured at 95˚C for 10 min, and cooled on ice. The denatured DNA was incubated with 5 μg of anti-5-methylcytosine antibody (Aviva Systems Biology, San Diego, CA) in 200 μl of IP buffer (10 mM sodium-phosphate, 140 mM NaCl, 0.05% Triton-X100 at pH7.0) for 3.5 hours at 4˚C. The antibody-bound DNA fragments were magnetically collected after incubation with Dynabeads M-280 (Life Technologies, Grand Island, NY) coated with anti-mouse IgG antibody for 3 hours at 4˚C. The beads were washed 5 times with 700 μl of IP buffer, resuspended in 250 μl of lysis buffer (50 mM TrisHCl at pH 8.0, 10 mM EDTA, 0.5% SDS) containing Proteinase K. After incubation at 55˚C for 30 min, DNA was recovered by phenol-chloroform extraction and subsequent ethanol precipitation.
|
| Label |
biotin
|
| Label protocol |
The immunoprecipitated DNA was amplified by WGA2 (Sigma Aldrich, Saint Louis, MO) to give a total of ~10 μg DNA, and then labeled with biotine-dUTP using GeneChip WT double stranded DNA terminal labeling kit (Affymetix, Santa Clara, CA).
|
| |
|
| |
| Hybridization protocol |
The labeled DNA was hybridized against a GeneChip Chromosome 21/22 2.0R Array (Affymetrix) at 45˚C for 16 hours. The array was washed and subsequently stained with Cy3-streptavidin in Fluidics Station 450 (Affymetrix) using GeneChip Hybridization, Wash, and Stain kit (Affymetrix).
|
| Scan protocol |
The fluorescent signal intensity at each probe spot was measured on GeneChip Scanner (Affymetrix).
|
| Description |
MeDIP-chip Technical Rep 1-3
|
| Data processing |
A log2-transformed ratio of signals obtained from MeDIP and control DNA was calculated for each probe by Tilling Analysis Software. The dataset was further processed by Baysian Tool for Methylation Analysis. The raw BATMAN scores were normalized (median=0, SD=1) for comparison.
.sgr contains normalized methylation score in 100-bp windows, which is generated by BATMAN software and perl script.
|
| |
|
| Submission date |
Feb 01, 2012 |
| Last update date |
Feb 02, 2012 |
| Contact name |
Kei Fukuda |
| E-mail(s) |
kei.fukuda@riken.jp
|
| Organization name |
RIKEN
|
| Lab |
Cellular memory
|
| Street address |
2-1 Hrosawa, Wakoshi
|
| City |
Saitama |
| State/province |
Japan |
| ZIP/Postal code |
351-0198 |
| Country |
Japan |
| |
|
| Platform ID |
GPL5715 |
| Series (1) |
| GSE35475 |
Identification of DNA methylation differences correlated with transcriptional divergence between humans and chimpanzees in chromosomes 21 and 22 |
|
| Supplementary file |
Size |
Download |
File type/resource |
| GSM869113_H3IP.CEL.gz |
21.4 Mb |
(ftp)(http) |
CEL |
| GSM869113_H3WG.CEL.gz |
20.2 Mb |
(ftp)(http) |
CEL |
| GSM869113_H4IP.CEL.gz |
21.0 Mb |
(ftp)(http) |
CEL |
| GSM869113_H4WG.CEL.gz |
21.0 Mb |
(ftp)(http) |
CEL |
| GSM869113_H5IP.CEL.gz |
20.7 Mb |
(ftp)(http) |
CEL |
| GSM869113_H5WG.CEL.gz |
20.5 Mb |
(ftp)(http) |
CEL |
| GSM869113_Human_meth_hg17.sgr.gz |
2.3 Mb |
(ftp)(http) |
SGR |
| Processed data provided as supplementary file |
|
|
|
|
 |
