|
| Status |
Public on Nov 30, 2017 |
| Title |
Scribbel-mut#1 |
| Sample type |
RNA |
| |
|
| Source name |
HepG2 cells expressing mutated Scribble
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HepG2
transfection: mutated Scribble
|
| Treatment protocol |
For the generation of cell lines stably expressing ScribWT and ScribP305L, HepG2 cells were transfected with 9 µg of pEGFP-ScribWT or pEGFP-ScribP305L on a 10 cm dish using Fugene HD (Promega, Mannheim, Germany). Selection with G418 (Geneticin, 1 mg/ml) was started 72 hours after transfection. Single clones were expanded and Scrib overexpression was analysed by Western blotting and real-time PCR.
|
| Growth protocol |
The human HCC cell lines were cultured in RPMI HepG2 supplemented with 10% fetal bo-vine serum and 1% penicillin/streptomycin at 37°C in a 5% CO2 atmosphere (Sigma-Aldrich, Taufkirchen, Germany). Cell lines were authenticated by STR-analysis (DSMZ, Braunschweig, Germany) and routinely checked for mycoplasma contamination.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For isolation of total RNA from cultured cells, the NucleoSpin RNA II kit (Machery-Nagel, Düren, Germany).
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol.
|
| |
|
| Hybridization protocol |
Hybridization (16h x 45°C) was processed according to the standard Affymetrix protocol.
|
| Scan protocol |
Affymetrix GeneArray Scanner3000
|
| Description |
Gene expression data from HepG2 cells expressing mutated Scribble
|
| Data processing |
The data were analyzed with a commercial software called JMP Genomics, version 7, from SAS. Gene expression profiling was performed using human transcriptome arrays 2.0 (HTA-2_0) from Affymetrix. A Custom CDF Version 20 with Entrez based gene definitions was used to annotate the arrays. The Raw fluorescence intensity values were normalized applying quantile normalization, RMA background correction and Medianpolish Probeset Summary
|
| |
|
| Submission date |
Jan 17, 2017 |
| Last update date |
Nov 30, 2017 |
| Contact name |
Carsten Sticht |
| Organization name |
University Heidelberg
|
| Department |
ZMF
|
| Street address |
Theodor-Kutzer-Ufer
|
| City |
Mannheim |
| ZIP/Postal code |
68169 |
| Country |
Germany |
| |
|
| Platform ID |
GPL22936 |
| Series (1) |
| GSE93742 |
Mislocalization of the cell polarity protein Scribble (Scrib) induces SPARC secretion in hepatocellular carcinoma |
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