|
| Status |
Public on Mar 06, 2009 |
| Title |
ribo deficient 24hr (EN 2) |
| Sample type |
RNA |
| |
|
| Source Name |
Caco-2
|
| Organism(s) |
Homo sapiens |
| Characteristics |
cell line: Caco-2
|
| Growth protocol |
Human colon epithelial (Caco-2) cells were cultured in DMEM or riboflavin-free DMEM (Invitrogen) containing 10% foetal calf serum (FCS) (Harlan Sera-Lab), 100 units/ml penicillin and 100 µg/ml streptomycin (Sigma). Dialysed FCS from the same batch was used where indicated. For clonogenic assays, 0.5 × 106 cells were treated as indicated to differing degrees of riboflavin depletion. After treatment, cells were harvested and plated out at 5,000 cells per well. After growth for 7 days, colonies were fixed, stained with methylene blue and counted
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen Rneasy RNA Extraction Kit was used for RNA extraction according to manufacturers' instructions
|
| Label |
biotin
|
| Label protocol |
Biotinylation of antisense RNA carried out according to Affymetrix protocol
|
| |
|
| Hybridization protocol |
15µg of cRNA molecules were heat fragmented and applied to the Human Genome U133 plus2 GeneChip in a hybridisation solution according to the Affymetrix protocol. Hybridisation took place overnight in a rotating hybridisation platform at 45°C for 16 hours. Post hybridisation stringency washing was done using the Affymetrix fluidics station again following the protocol outlined in the Affymetrix instructions
|
| Scan protocol |
the GeneChips were scanned in the Affymetrix GCS3000 scanner and the resultant image processed using the GCOS software package to prepare the .CEL files
|
| Description |
Ribo deficient cells after 24 hour treatment
Gene expression data from human colonic epithelial cells.
|
| Data processing |
The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method. The trimmed mean target intensity of each array was arbitrarily set to 100
|
| |
|
| Submission date |
Mar 05, 2009 |
| Contact name |
Paul Heath |
| E-mail(s) |
p.heath@sheffield.ac.uk
|
| Phone |
+44 114 2712473
|
| Organization name |
University of Sheffield
|
| Department |
Academic Neurology Unit
|
| Street address |
Medical School, Beech Hill Road
|
| City |
Sheffield |
| ZIP/Postal code |
S10 2RX |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL570 |
| Series (1) |
| GSE15132 |
Riboflavin depletion impairs cell proliferation in intestinal cells: Identification of mechanisms and consequences |
|
