|
| Status |
Public on Apr 17, 2012 |
| Title |
iPS actinomycin D 120 min rep 3 |
| Sample type |
RNA |
| |
|
| Source name |
iPSCs with actinomycin D, time 120 min
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: iPS cells (induced pluripotent stem cells)
treatment: actinomycin D
time: 120 min
|
| Treatment protocol |
To inhibit transcription, cells were treated with 5µg/ml actinomycin D (Invitrogen) for 20 minutes.
|
| Growth protocol |
Human foreskin fibroblasts (HFFs) and genetically identical in Duced pluripotent stem (iPS) cells were purchased from System Biosciences. The HFFs were cultured in DMEM medium supplemented with 10% FBS, 2mM L-glutamine, 100µM nonessential amino acids, 50U/ml penicillin and 50µg/ml streptomycin. The iPS cells were cultured in mTeSR®1 medium (Stemcell Technologies Inc) supplemented with 50U/ml penicillin and 50µg/ml streptomycin on plates coated with 0.3mg/ml Matrigel™ Basement Membrane Matrix Growth Factor Reduced (BD Biosciences). Cells were incubated at 37°C with 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were collected in Trizol® (Invitrogen) at 0, 15, 30, 160, 120 and 240 minutes after transcription inhibition. Total RNA was isolated according to the manufacturer’s protocol. RNA quality and concentration were analyzed using a NanoDrop 2000c spectrophotometer (Thermo Fisher Scientific) and Bioanalyzer (Agilent).
|
| Label |
biotin
|
| Label protocol |
Biotinylated DNA probes were prepared according to Whole Transcript Sense Target Labeling Manual (Affymetrix)
|
| |
|
| Hybridization protocol |
Labeled probes were hybridized to GeneChip Human Gene 1.0 ST Arrays according to GeneChip® WT Terminal Labeling and Hybridization User Manual provided with the WT Expression Kit (Affymetrix)
|
| Scan protocol |
Labeled hybridized arrays were washed and stained using the Fluidics Station 450 prior to scanning on the GeneChip Scanner 3000 7G according to GeneChip® Expression Wash, Stain and Scan Manual (Affymetrix)
|
| Description |
This is sample was collected at 120 min after actinomycin D treatment
|
| Data processing |
All the probe sets were applied median normalization by Affymetrix Power Tools (APT) with ‘no adjustment’ option and were normalized to the 5th percentile value of all probe sets on the same array.
|
| |
|
| Submission date |
Nov 02, 2011 |
| Last update date |
Apr 17, 2012 |
| Contact name |
Ju Youn Lee |
| Organization name |
UMDNJ-New Jersey Medical School
|
| Department |
Biochemistry
|
| Street address |
185 South Orange Ave. MSB-E501
|
| City |
Newark |
| State/province |
NJ |
| ZIP/Postal code |
07101 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE33417 |
Global analysis of mRNA decay in induced pluripotent stem cells |
|
