|
| Status |
Public on May 04, 2015 |
| Title |
Hepatocellular carcinoma tissue_50t |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
MeDIP of hepatocellular carcinoma tissue
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Hepatocellular carcinoma
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
After surgical excision, tissue samples for nucleic acids extraction were immediately sliced into aliquots of about 100 mg, snap-frozen in liquid nitrogen and stored at -80°C until use. Aliquots for DNA extraction were homogenized in 2ml of chilled NaCl 0.9% w/v; cell lysis was achieved by using Nonidet P40 (Sigma-Aldrich, St. Louis, MO, USA) 0.1% and lysis solution (NaCl 100 mM, EDTA 25 mM, SDS 1.6 %, pH 8). Samples were treated with proteinase K/RNase and DNA extracted with a standard phenol/chloroform procedure. The methylated DNA immunoprecipitation (MeDIP) was performed with MeDIP kit™ by Diagenode (Liège, Belgium). IP and INPUT samples were amplified by GenomePlex Complete Genome Amplification (WGA) kit (Sigma-Aldrich, St. Louis, MO, USA) following the producer’s protocol.
|
| Label |
Cy5
|
| Label protocol |
One and a half μg of IP and 1.5 μg INPUT samples were labelled with Cy5 and Cy3 respectively by Dual-Color DNA Labeling Kit (NimbleGen-Roche, Madison, WI, USA).
|
| |
|
| Channel 2 |
| Source name |
INPUT DNA from hepatocellular carcinoma tissue
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Hepatocellular carcinoma
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
After surgical excision, tissue samples for nucleic acids extraction were immediately sliced into aliquots of about 100 mg, snap-frozen in liquid nitrogen and stored at -80°C until use. Aliquots for DNA extraction were homogenized in 2ml of chilled NaCl 0.9% w/v; cell lysis was achieved by using Nonidet P40 (Sigma-Aldrich, St. Louis, MO, USA) 0.1% and lysis solution (NaCl 100 mM, EDTA 25 mM, SDS 1.6 %, pH 8). Samples were treated with proteinase K/RNase and DNA extracted with a standard phenol/chloroform procedure. The methylated DNA immunoprecipitation (MeDIP) was performed with MeDIP kit™ by Diagenode (Liège, Belgium). IP and INPUT samples were amplified by GenomePlex Complete Genome Amplification (WGA) kit (Sigma-Aldrich, St. Louis, MO, USA) following the producer’s protocol.
|
| Label |
Cy3
|
| Label protocol |
One and a half μg of IP and 1.5 μg INPUT samples were labelled with Cy5 and Cy3 respectively by Dual-Color DNA Labeling Kit (NimbleGen-Roche, Madison, WI, USA).
|
| |
|
| |
| Hybridization protocol |
Samples were hybridized on the Human DNA Methylation 3x720K CpG Island Plus RefSeq Promoter Array (NimbleGen-Roche, Madison, WI, USA) following manufacturer instructions
|
| Scan protocol |
Microarrays were scanned with Axon GenePix 4400A microarray scanner setting laser power to 100%. PMT gain was set in order to obtain about 1e-5 normalized counts at 65.000 intensity level for both channels.
|
| Description |
MeDIP-chip hepatocellular carcinoma tissue
|
| Data processing |
Data were quantile normalized and logged fold change of immunoprecipitated sample (IP) over control (INPUT) sample was calculated
|
| |
|
| Submission date |
Jul 09, 2014 |
| Last update date |
May 05, 2015 |
| Contact name |
Alberto Ferrarini |
| E-mail(s) |
alberto.ferrarini@univr.it
|
| Phone |
+39-045-802-7058
|
| Organization name |
University of Verona
|
| Department |
Scientific and Technological Department
|
| Lab |
Plant Functional Genomics Centre
|
| Street address |
Strada le Grazie, 15
|
| City |
Verona |
| State/province |
Veneto |
| ZIP/Postal code |
37134 |
| Country |
Italy |
| |
|
| Platform ID |
GPL15160 |
| Series (2) |
| GSE59260 |
DNA methylation profiles in alcohol-associated hepatocellular carcinoma |
| GSE59261 |
Expression and DNA methylation profiles in alcohol-associated hepatocellular carcinoma |
|
