|
| Status |
Public on Nov 26, 2013 |
| Title |
JNK1,2 KO MEFs_d6_3 |
| Sample type |
RNA |
| |
|
| Source name |
JNK1,2-deficient MEFs
|
| Organism |
Mus musculus |
| Characteristics |
strain background: BL6/129/Sv
genotype/variation: jnk1-/-jnk2-/-
tissue: mouse embryo
developmental stage: E10.0
cell type: immortalized 3T3 murine embryonic fibroblasts (MEFs)
|
| Treatment protocol |
Murine embryonic fibroblasts were isolated from JNK1,2-deficient and wildtype E10.0 embryos and immortalized following the 3T3 protocol. Cells were published previously in Sabapathy et al. (2004) Mol Cell 15:713-25.
|
| Growth protocol |
Cells were cultured at 37°C and 8% CO2 in a humidified atmosphere on cell culture inserts (BD Biosciences) in FAD medium in absence (day 0) or presence (days 2, 4, 6) of NHEK. The FAD medium is composed of DMEM:Ham´s F12 (3:1) without calcium chloride supplemented with 5 % calcium-depleted FBS, 2 mM L-glutamine, 100 U ml-1 penicillin, 0.1 mg ml-1 streptomycin, 5 µg/ml insulin, 0.4 µg/ml hydrocortisone, 10-10 M cholera toxin, 24 ng/ml adenine, 1 ng/ml hEGF, 200 µM CaCl2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with RLT buffer provided in the QIAGEN RNeasy mini kit, followed by clean-up and DNase I treatment with QIAGEN RNeasy mini kit in accordance with the prescribed protocol provided with the kit. Quality control of all samples was performed by measuring quality and concentration of RNA with Nanodrop ND-1000 and Agilent 2100 Bioanalyzer.
|
| Label |
biotin
|
| Label protocol |
500 ng of total RNA were reverse transcribed into cDNA, which was further amplified to generate biotin-labeled cRNA by in vitro transcription using the Illumina® Total Prep™ RNA Amplification Kit (Life Technologies). Following column-based purification, eluted cRNA was re-assessed for quality and quantity using the RNA 6000 Nano Kit on an Agilent 2100 Bioanalyzer and the NanoDrop ND-1000 spectrophotometer, respectively.
|
| |
|
| Hybridization protocol |
Standard Illumina hybridization protocol
|
| Scan protocol |
The arrays were scanned using an iScan array scanner (Illumina Inc.)
|
| Description |
JNK1,2-deficient MEFs post triad 3; rep 3; 1 cell culture insert was used for this sample
|
| Data processing |
The data were normalised using quantile normalisation according to Smyth GK, Speed T (2003) Methods 31:265-73.
|
| |
|
| Submission date |
Jul 09, 2013 |
| Last update date |
Nov 26, 2013 |
| Contact name |
Peter Angel |
| E-mail(s) |
p.angel@dkfz-heidelberg.de
|
| Phone |
00496221424570
|
| Organization name |
German Cancer Research Center
|
| Department |
Signal Transduction and Growth Control
|
| Street address |
Im Neuenheimer Feld 280
|
| City |
Heidelberg |
| ZIP/Postal code |
69124 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6887 |
| Series (2) |
| GSE48612 |
Genome-wide analysis of gene expression in jnk1-/-jnk2-/- immortalized mouse embryonic fibroblasts compared to wildtype counterparts co-cultivated with differentiating keratinocytes |
| GSE48614 |
Array_data_MEF_Kera_Coculture |
|
