|
| Status |
Public on Dec 31, 2016 |
| Title |
patient 47, tumoral sample1, array 2 |
| Sample type |
RNA |
| |
|
| Source name |
colon adenocarcinoma
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: colon adenocarcinoma
tissue preparation: frozen sample
anatomical site: left colon
gender: male
patient age: 77
tumor stage: tumor stage 2
|
| Growth protocol |
Caco-2 and HT29 cell lines have been cultured in D-MEM medium (Dulbecco's Modified Eagle Medium 1X; GIBCO, COD:31965-023 containing 4.5g/L of D-glucose), supplemented with 10% FBS (Fetal Bovine Serum) and 100 U/ml of antibiotics (Penicillin-Streptomycin). HCT116 cell line was cultured in McCoy 5A+ (McCoy’s 5A Medium 1X GIBCO, COD: 36600021 containing 3 g/L of D-glucose) supplemented with 10%FBS (Fetal Bovine Serum), 1.5 mM L-glutamine and 2200 mg/L of sodium bicarbonate. Cell lines were incubated at 37 °C in humidified atmosphere with 5% of CO2 and 95% of air. The culture medium was changed twice a week, then RNA was extracted using RNeasy Mini Kit- (Qiagen, Milan-Italy).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA (100 ng) was extracted from frozen tissue and cell cultures using RNeasy Mini Kit (Qiagen, Milan - Italy) according to the manufacturer’s instructions. The concentration and the quality of RNA were determined using a ND-1000 spectrophotometer (NanoDrop, Thermo Scientific, USA).
|
| Label |
biotin
|
| Label protocol |
Biotinylated fragmented sense strand cDNA were prepared according to the protocol supplied by the Affymetrix for Human Transcriptome Array 2.0 (HTA).
|
| |
|
| Hybridization protocol |
Fragmented and labeled target (Sense Strand-cDNA 5.2-5.5 µg) was used for hybridization to the “GeneChip Human Transcriptome Array 2.0” according to the protocol supplied by the manufacturer was used for hybridization step. The hybridization has been performed in an hybridization oven at 45°C for 16 hours with rotation at 60 rpm.
|
| Scan protocol |
Scanning was performed using the Affymetrix Scanner GCS3000 7G.
|
| Data processing |
Data analysis was performed using Affymetrix® Expression Console™ software (v.1.4). Expression level analysis was obtained using the normalization method based on the processing algorithm Robust Multi-array Average (RMA).
|
| |
|
| Submission date |
Sep 23, 2015 |
| Last update date |
Dec 31, 2016 |
| Contact name |
Daniele F Condorelli |
| E-mail(s) |
daniele.condorelli@unict.it
|
| Organization name |
University of Catania
|
| Department |
Biomedical and Biotechnological Sciences
|
| Lab |
Medical Biochemistry
|
| Street address |
Via S. Sofia 64
|
| City |
Catania |
| State/province |
Italy |
| ZIP/Postal code |
95125 |
| Country |
Italy |
| |
|
| Platform ID |
GPL17586 |
| Series (1) |
| GSE73360 |
Human Transcriptome Array 2.0 (HTA) in matched colorectal cancer and normal colonic mucosa |
|
