|
| Status |
Public on Jan 01, 2018 |
| Title |
bladder 10y |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Frozen tissue biopsy
|
| Organism |
Homo sapiens |
| Characteristics |
disease state: Urothelial cancer
tissue: tumor (Frozen tissue biopsy)
location: bladder
timepoint: 10y
|
| Treatment protocol |
None
|
| Growth protocol |
Tumor samples from the patient from different locations and different timepoints with urothelial carcinoma were either snap-frozen and maintaned at -80 degrees C or FFPE archived 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 |
reference: Normal female genome
|
| Treatment protocol |
None
|
| Growth protocol |
Tumor samples from the patient from different locations and different timepoints with urothelial carcinoma were either snap-frozen and maintaned at -80 degrees C or FFPE archived 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 visualized and analyzed using DNA Analytics 4.0.85 and the ADM2 step gram algorithm.
For normalization, we applied GC-correction and diploid peak-centralization using the Agilent Workbench 7.0.4.0.
|
| |
|
| Submission date |
Dec 01, 2016 |
| Last update date |
Jan 01, 2018 |
| Contact name |
David Christian Müller |
| E-mail(s) |
david.mueller@usb.ch
|
| Organization name |
University Hospital Basel
|
| Department |
Institute for Pathology
|
| Street address |
Schönbeinstrasse 40
|
| City |
Basel |
| ZIP/Postal code |
4031 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL10123 |
| Series (1) |
| GSE90778 |
Array-CGH data of aneuploid tumor populations of different urothelial cancer sites in one patient |
|
