|
| Status |
Public on Dec 31, 2010 |
| Title |
E15 testis sample1 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Cy3, total input sonicated DNA
|
| Organism |
Mus musculus |
| Characteristics |
tissue: testis
developmental stage: E15
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total genomic DNA was extracted using Qiagen DNeasy kit (Qiagen) according to the manufacture's instruction. DNA was sonicated to give fragments between 200bp and 1000bp.
|
| Label |
Cy3
|
| Label protocol |
Labeling performed according to standard procedures by Nimblegen
|
| |
|
| Channel 2 |
| Source name |
Cy5, methylated DNA enriched by MeDIP
|
| Organism |
Mus musculus |
| Characteristics |
tissue: testis
developmental stage: E15
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Sonicated total genomic DNA was digested with MseI (TTAA) (New England Biolabs) to produce small fragments (200 - 1,000bp) while keeping CpG islands intact. Fragmented DNA was heat denatured to produce single-stranded DNA, and a portion of the denatured DNA stored as control (input) DNA. Monoclonal mouse anti 5-methyl cytidine (Eurogentec, Reference ID: BI-MECY-0500; batch#: 080128) was used to immunoprecipitate methylated DNA fragments. The immune complexes were captured with Protein A agarose beads (Invitrogen). Complexes were washed to remove nonspecifically bound material. MeDIP DNA was obtained by elution of bound complexes and ethanol precipitation.
|
| Label |
Cy5
|
| Label protocol |
Labeling performed according to standard procedures by Nimblegen
|
| |
|
| |
| Hybridization protocol |
Hybridization performed according to standard procedures by Nimblegen
|
| Scan protocol |
Scanning performed according to standard procedures by Nimblegen using NimbleScan.
|
| Data processing |
Arrays were processed using Nimblegen's standard protocol for Nimblescan 2.4 ChIP data extraction. VALUE is the P score. To calculate the P score, the ratio of the input signals for input DNA and the MeDIP DNA from the same sample that were co-hybridized to the array is first computed and scaled to center the ratio data around zero. Scaling is performed by subtracting the bi-weight mean for the log2-ratio values for all features on the array from each log2-ratio value. A fixed-length window (750bp) is then placed around each consecutive probe and the one-sided Kolmogorov-Smirnov (KS) test is applied to determine whether the probes are drawn from a significantly more positive distribution of intensity log-ratios than those in the rest of the array. The resulting score for each probe, called P score, is the -log10 p-value from the windowed KS test around that probe. The identification of methylation peaks is done by searching for at least 2 consecutive probes above a P score of 2. Peaks within 500bp of each other are merged.
|
| |
|
| Submission date |
Apr 20, 2010 |
| Last update date |
Dec 31, 2010 |
| Contact name |
Ping Liang |
| E-mail(s) |
[email protected]
|
| Phone |
1(905)6885550-5922
|
| Organization name |
Brock University
|
| Department |
Department of Biological Sciences
|
| Street address |
500 Glenridge Ave
|
| City |
St. Catharines |
| State/province |
Ontario |
| ZIP/Postal code |
L2S3A1 |
| Country |
Canada |
| |
|
| Platform ID |
GPL7060 |
| Series (1) |
| GSE21415 |
Genome-wide survey reveals dynamic widespread tissue-specific changes in DNA methylation during development |
|
