|
| Status |
Public on Oct 30, 2018 |
| Title |
454199_A02 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
MeDIP DNA_EPN - 150ppm - Methylation
|
| Organism |
Rattus norvegicus |
| Characteristics |
Sex: Female
tissue: hippocampus
strain: Long-Evans
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted from hippocampus tissue samples using PureGene DNA isolation kits.
|
| Label |
Cy5
|
| Label protocol |
Experimental (IP) samples were labeled with Cy5 and control (input) samples with Cy3.
|
| |
|
| Channel 2 |
| Source name |
Input DNA_EPN - 150ppm - Methylation
|
| Organism |
Rattus norvegicus |
| Characteristics |
Sex: Female
tissue: hippocampus
strain: Long-Evans
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted from hippocampus tissue samples using PureGene DNA isolation kits.
|
| Label |
Cy3
|
| Label protocol |
Experimental (IP) samples were labeled with Cy5 and control (input) samples with Cy3.
|
| |
|
| |
| Hybridization protocol |
The immunoprecipitated CpG-methylated DNA (test, experimental samples) and the untreated, sonicated DNA (input DNA) were differentially labeled and hybridized to a single array as a two-color experiment (Rat DNA Methylation 3 × 720 K CpG Island Plus RefSeq Promoter Array (GPL 18610) for 20 h at 42°C, followed by washing with the Nimblegen Wash Kit.
|
| Scan protocol |
Scanning was performed with 100% gain and double pass for optimal sensitivity and scanning accuracy.
|
| Description |
EPN - 150ppm - Methylation
MeDIP_Hippocampus
|
| Data processing |
Raw data were extracted as pair files by NimbleScan software. The intensity ratio of immunoprecipitated to control DNA was plotted versus genomic position to identify regions where increased signal (i.e. DNA fragment enrichment) was present relative to the control sample. The R/Bioconductor package Ringo was utilized to extract the green (Cy3, 532nm, associated to total DNA) and red (Cy5, 635nm, associated to MeDIP fraction) channels from the provided pair files. Normalization of the probe intensities was carried out by applying the Nimblegen algorithm, following the recommendation of Adriaens et al. for ChIP-on-chip and DNA methylation microarrays (Adriaens et al., 2012).
Signals from the two channels were then summarized by calculating the log2 ratio of the MeDIP and input channels. Being log2-ratios, a value of 1 corresponds to a two-fold enrichment of the MeDIP signal relative to total DNA, while a value of -1 is interpreted as a MeDIP signal two times smaller than the input channel. The normalized data for the control "RANDOM" probes are available on the series record.
|
| |
|
| Submission date |
Oct 30, 2017 |
| Last update date |
Oct 30, 2018 |
| Contact name |
Jay S Schneider |
| E-mail(s) |
jay.schneider@Jefferson.edu
|
| Organization name |
Thomas Jefferson University
|
| Department |
Anatomy,Pathology and Cell biology
|
| Lab |
Room511
|
| Street address |
1020 Locust St
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19107 |
| Country |
USA |
| |
|
| Platform ID |
GPL16994 |
| Series (1) |
| GSE106323 |
Hippocampal methylome of rats exposed to lead at different developmental stages |
|
