|
| Status |
Public on Mar 31, 2007 |
| Title |
Melanoma cell line D24 |
| Sample type |
RNA |
| |
|
| Source name |
Melanoma cell line D24
|
| Organism |
Homo sapiens |
| Characteristics |
Melanoma cell line established from metastatic melanoma
|
| Biomaterial provider |
Dr C. Schmidt
|
| Growth protocol |
Melanoma cell lines were cultured in RPMI 1640 in the presence of 10% fetal bovine serum.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extractions were performed using Qiagen RNeasy Midi-kits as per the manufacturer’s instructions. RNA quality and concentration were assessed using a 2100 BioAnalyser (Agilent Technologies).
|
| Label |
biotin
|
| Label protocol |
For each sample, 5 μg of total RNA were used for target preparation using Eukaryotic One-cycle Target Labelling and Control Reagents kits according to the manufacturer’s instructions (Affymetrix). First-strand complementary DNA (cDNA) was synthesized using a T7-Oligo(dT) primer followed by second-strand cDNA synthesis from these samples using T4 polymerase. An in-vitro transcription reaction was then performed to produce biotin-labelled complementary RNA (cRNA).
|
| |
|
| Hybridization protocol |
This cRNA was fragmented and 15 μg were hybridized to an Affymetrix Human Genome U133 Plus 2.0 Array in an Affymetrix Hybridization Oven 640 for 16 h at 45oC. Chips were then washed and post-stained with streptavidin phycoerythrin and normal goat IgG solutions on an Affymetrix Fluidics Station 450.
|
| Scan protocol |
Chips were scanned on an Affymetrix GeneChip Scanner 3000, and raw image data files obtained using Affymetrix GeneChip Operating Software (GCOS Version 1.2).
|
| Description |
For each sample, 5 μg of total RNA were used for target preparation using Eukaryotic One-cycle Target Labelling and Control Reagents kits according to the manufacturer’s instructions (Affymetrix). First-strand complementary DNA (cDNA) was synthesized using a T7-Oligo(dT) primer followed by second-strand cDNA synthesis from these samples using T4 polymerase. An in-vitro transcription reaction was then performed to produce biotin-labelled complementary RNA (cRNA). This cRNA was fragmented and 15 μg were hybridized to an Affymetrix Human Genome U133 Plus 2.0 Array in an Affymetrix Hybridization Oven 640 for 16 h at 45oC. Chips were then washed and post-stained with streptavidin phycoerythrin and normal goat IgG solutions on an Affymetrix Fluidics Station 450. Chips were scanned on an Affymetrix GeneChip Scanner 3000, and raw image data files obtained using Affymetrix GeneChip Operating Software (GCOS Version 1.2). Relative expression values were generated for each probe set using the Affymetrix PLIER algorithm in ArrayAssist® Version 4.20 (Stratagene) and further log transformed to base 2. The resulting data were imported into GeneSpring GX v7.3 (Agilent Technologies). Data values less than 0.1 were set to 0.1 and the logarithm of data values were used as expression values. The expression values were normalized in two steps. First, values were centralized sample by sample, such that the median expression value was zero for each sample. Second, expression values were centralized gene by gene such that the median expression value was zero for each probe set.
|
| Data processing |
Relative expression values were generated for each probe set using the Affymetrix PLIER algorithm in ArrayAssist® Version 4.20 (Stratagene) and further log transformed to base 2. The resulting data were imported into GeneSpring GX v7.3 (Agilent Technologies). Data values less than 0.1 were set to 0.1 and the logarithm of data values were used as expression values. The expression values were normalized in two steps. First, values were centralized sample by sample, such that the median expression value was zero for each sample. Second, expression values were centralized gene by gene such that the median expression value was zero for each probe set.
|
| |
|
| Submission date |
Feb 19, 2007 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Leisl Packer |
| URL |
http://www.qimr.edu.au
|
| Organization name |
Queensland Institute of Medical Research
|
| Department |
Cancer and Cell Biology
|
| Lab |
Oncogenomics Laboratory
|
| Street address |
300 Herston Rd
|
| City |
Herston |
| State/province |
QLD |
| ZIP/Postal code |
4006 |
| Country |
Australia |
| |
|
| Platform ID |
GPL570 |
| Series (2) |
| GSE7127 |
63 Melanoma cell lines |
| GSE7152 |
Melanoma cell lines involved in the p14ARF genotype analysis |
|
| Relations |
| Affiliated with |
GSM700981 |
| Affiliated with |
GSM804316 |
| Affiliated with |
GSM804317 |
| Reanalyzed by |
GSE64985 |
| Reanalyzed by |
GSE119087 |
