|
| Status |
Public on Jul 04, 2011 |
| Title |
P1018R |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Frozen tissue biopsy
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Pancreatic Ductal Adenocarcinoma
|
| Growth protocol |
Tumor samples from patients with either pancreatic ductal adenocarcinoma, adrenocortical carcinoma, or prostate carcinoma were flash-frozen and maintaned at -80 degrees C until processed for flow sorting
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
25-100 mg of tumor was thawed then minced in the presence of NST and DAPI. Each sample was then mechanically sheared then filtered through a 30 micron mesh filter prior to sorting. All sorting was done with an Influx Flow Cytometer equiped with a UV laser. DNA was extracted from each sorted fraction using Qiagen DNA micro kits. All phi29 amplifications were done with a 100ng aliquot of either sample or reference genomic DNA using the GenomiPhi kit (G.E Health Care).
|
| Label |
Cy5
|
| Label protocol |
A 1 microgram aliquot of each sample and reference DNA was digested with DNAse 1 then labeled with a fluorescent nucleotide (Cy5 for tumor and Cy3 for normal reference) using the BioPrime labeling kit (Invitrogen)according to manufacturer's protocol.
|
| |
|
| Channel 2 |
| Source name |
Reference pooled 46XX
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: pooled normal (46,XX) reference (Promega)
|
| Growth protocol |
Tumor samples from patients with either pancreatic ductal adenocarcinoma, adrenocortical carcinoma, or prostate carcinoma were flash-frozen and maintaned at -80 degrees C until processed for flow sorting
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
25-100 mg of tumor was thawed then minced in the presence of NST and DAPI. Each sample was then mechanically sheared then filtered through a 30 micron mesh filter prior to sorting. All sorting was done with an Influx Flow Cytometer equiped with a UV laser. DNA was extracted from each sorted fraction using Qiagen DNA micro kits. All phi29 amplifications were done with a 100ng aliquot of either sample or reference genomic DNA using the GenomiPhi kit (G.E Health Care).
|
| Label |
Cy3
|
| Label protocol |
A 1 microgram aliquot of each sample and reference DNA was digested with DNAse 1 then labeled with a fluorescent nucleotide (Cy5 for tumor and Cy3 for normal reference) using the BioPrime labeling kit (Invitrogen)according to manufacturer's protocol.
|
| |
|
| |
| Hybridization protocol |
Labeled DNAs were hybridized in an ozone free environment in a rotisserie oven at 20 rpm for 40 hours then washed according to array supplier's (Agilent)protocol
|
| Scan protocol |
All arrays were scanned using an Agilent 2565C Scanner and default settings for CGH.
|
| Data processing |
Data was extracted from the TIFF files using Agilent FE 10.5. The data quality was assessed using the QC Report output in F.E. 10.5. All arrays that passed the experimental Q.C. were then visulaized and analyzed using DNA Analytics 4.0.85 and the ADM2 step gram algorithm.
|
| |
|
| Submission date |
May 04, 2010 |
| Last update date |
Jul 04, 2011 |
| Contact name |
Michael Thomas Barrett |
| E-mail(s) |
barrett.michael@mayo.edu
|
| Phone |
480-301-6736
|
| Organization name |
Mayo Clinic Arizona
|
| Department |
Molecular Pharmacology and Experimental Therapeutics
|
| Street address |
13400 East Shea Boulevard
|
| City |
Scottsdale |
| State/province |
AZ |
| ZIP/Postal code |
85259 |
| Country |
USA |
| |
|
| Platform ID |
GPL4091 |
| Series (1) |
| GSE21660 |
Advancing a Clinically Relevant Perspective of the Clonal Nature of Cancer |
|
