|
| Status |
Public on Aug 06, 2011 |
| Title |
Fibroblast_HELP |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
LM-PCR products from HpaII digested DNA
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Foreskin fibroblast
digestive enzyme: HpaII
|
| Growth protocol |
The cells were grown in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% Fetal Bovine Serum, 2 mM Glutamine and Penicillin-Streptomycin (Invitrogen) in a 37°C incubator with 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was isolated using proteinase K digestion, phenol-chloroform extraction, dialysis against 0.2X SSC and concentration by surrounding the dialysis bag with PEG 20,000 to reduce water content by osmosis. The quality of the DNA was checked by gel electrophoresis and spectrophotometry.
|
| Label |
Cy5
|
| Label protocol |
The HELP samples were labeled for microarray analysis as described previously[Selzer, Genes Chromosomes Cancer 44:305–319] using Cy3 or Cy5-conjugated oligonucleotides and random primers.
|
| |
|
| Channel 2 |
| Source name |
LM-PCR products
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Foreskin fibroblast
digestive enzyme: Msp 1
|
| Growth protocol |
The cells were grown in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% Fetal Bovine Serum, 2 mM Glutamine and Penicillin-Streptomycin (Invitrogen) in a 37°C incubator with 5% CO2.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was isolated using proteinase K digestion, phenol-chloroform extraction, dialysis against 0.2X SSC and concentration by surrounding the dialysis bag with PEG 20,000 to reduce water content by osmosis. The quality of the DNA was checked by gel electrophoresis and spectrophotometry.
|
| Label |
Cy3
|
| Label protocol |
The HELP samples were labeled for microarray analysis as described previously[Selzer, Genes Chromosomes Cancer 44:305–319] using Cy3 or Cy5-conjugated oligonucleotides and random primers.
|
| |
|
| |
| Hybridization protocol |
The HpaII and MspI representations were co-hybridized to the microarray and scanned to quantify the 532 and 635 nm fluorescence at each oligonucleotide on the microarray.
|
| Scan protocol |
see manufacturer's web site www.nimblegen.com
|
| Data processing |
HELP assay data analysis was performed using our published pipeline (Thompson, Reimers, Khulan, Gissot, Richmond, Chen, Zheng, Kim and Greally 2008), an open source resource available through BioConductor (HELP package)
|
| |
|
| Submission date |
Feb 24, 2011 |
| Last update date |
Apr 11, 2012 |
| Contact name |
John Greally |
| E-mail(s) |
[email protected]
|
| Phone |
7186781234
|
| Organization name |
Albert Einstein College of Medicine
|
| Department |
Genetics
|
| Street address |
1301 Morris Park Avenue
|
| City |
Bronx |
| State/province |
NY |
| ZIP/Postal code |
10461 |
| Country |
USA |
| |
|
| Platform ID |
GPL13223 |
| Series (2) |
| GSE27537 |
LATE-REPLICATING HETEROCHROMATIN IS CHARACTERISED BY DECREASED CYTOSINE METHYLATION IN THE HUMAN GENOME (HELP assay) |
| GSE27538 |
LATE-REPLICATING HETEROCHROMATIN IS CHARACTERISED BY DECREASED CYTOSINE METHYLATION IN THE HUMAN GENOME |
|
