|
| Status |
Public on Apr 04, 2016 |
| Title |
buccal cells_29_rep_1 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
buccal cells, control
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: buccal cells
disease status: control
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The samples were first digested using proteinase K (20mg/ml) and lysis buffer mixture at 55°C overnight. A mixture of phenol, chloroform, and iso-amyl alcohol (in ratios of 25:24:1) was used to remove the proteins, followed by two times chloroform wash, precipitation of DNA using cold isopropanol, and a final wash using 70% ethanol. After the extraction, the DNA samples were suspended in 0.1M Tris-EDTA buffer and were kept in -80°C for storage prior to use. The purity and the concentrations of DNA were checked using a UV spectrophotometer.
|
| Label |
Cy3
|
| Label protocol |
1 μg of purified enriched PCR products were fluorescently labelled with Cy5 (Roche) for the reference and with Cy3 (Roche) for the sample, as previously described (Schumacher, A., et al. (2006). Microarray-based DNA methylation profiling: technology and applications. Nucleic acids research 34, 528-542.). Each sample was independently enriched and hybridized twice to produce two technical replicates.
|
| |
|
| Channel 2 |
| Source name |
white blood cells, common reference pool
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: white blood cells
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The samples were first digested using proteinase K (20mg/ml) and lysis buffer mixture at 55°C overnight. A mixture of phenol, chloroform, and iso-amyl alcohol (in ratios of 25:24:1) was used to remove the proteins, followed by two times chloroform wash, precipitation of DNA using cold isopropanol, and a final wash using 70% ethanol. After the extraction, the DNA samples were suspended in 0.1M Tris-EDTA buffer and were kept in -80°C for storage prior to use. The purity and the concentrations of DNA were checked using a UV spectrophotometer.
|
| Label |
Cy5
|
| Label protocol |
1 μg of purified enriched PCR products were fluorescently labelled with Cy5 (Roche) for the reference and with Cy3 (Roche) for the sample, as previously described (Schumacher, A., et al. (2006). Microarray-based DNA methylation profiling: technology and applications. Nucleic acids research 34, 528-542.). Each sample was independently enriched and hybridized twice to produce two technical replicates.
|
| |
|
| |
| Hybridization protocol |
The enriched PCR products were hybridized overnight at 48°C onto the 12.1k human CpG island microarray (Microarray Facility, University Health Network, Toronto) as previously described (Schumacher, A., et al. (2006). Microarray-based DNA methylation profiling: technology and applications. Nucleic acids research 34, 528-542.) .
|
| Scan protocol |
The arrays were scanned using the Genepix 4000B scanner and Genepix Pro 6.0 (Axon Instruments).
|
| Description |
Duke Twins Study of Memory in Aging twin15b vs. common reference pool
BUCCAL_30A
|
| Data processing |
Linear Models for Microarray Data (LIMMA) package (Bioconductor) for R was used for background correction, normalization, and to extract the signal intensities. Briefly, normexp was used for background correction, loess was used for within-array normalization, and Rquantile was used for between-array normalization.
|
| |
|
| Submission date |
Sep 09, 2014 |
| Last update date |
Apr 04, 2016 |
| Contact name |
Art Petronis |
| Organization name |
Centre for Addiction and Mental Health
|
| Department |
Campbell Family Mental Health Research Institute
|
| Lab |
Krembil Family Epigenetics Laboratory
|
| Street address |
250 College St
|
| City |
Toronto |
| State/province |
Ontario |
| ZIP/Postal code |
M5T1R8 |
| Country |
Canada |
| |
|
| Platform ID |
GPL2040 |
| Series (1) |
| GSE61242 |
Epigenetic assimilation in the aging human brain |
|
