|
| Status |
Public on Mar 16, 2010 |
| Title |
HP1-RNA-4, expression |
| Sample type |
RNA |
| |
|
| Source name |
Kc cells treated with HP1 dsRNA
|
| Organism |
Drosophila melanogaster |
| Characteristics |
gender: female
|
| Treatment protocol |
Double stranded RNA (dsRNA) was prepared from a PCR product spanning the entire HP1 coding region, generated with primers containing a T7 RNA Polymerase binding site using the MEGASCRIPT T7 In Vitro Transcription Kit (Ambion, Cat.No. AM1334). The RNA was purified and heated to 70○C for 10 minutes and slowly cooled down to room temperature for about 30 minutes to enhance annealing. 50μg of dsRNA was added to 106 cells every 2nd day and 8 days after initial addition of dsRNA cells were harvested and the efficiency of HP1 reduction was estimated by western blot analysis using a monoclonal α-HP1 antibody.
|
| Growth protocol |
Drosophila Kc cells were kept in HyQ-SFX (Hyclone).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Drosophila Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
|
| Scan protocol |
GeneChips were scanned on an Affymetrix Scanner 3000 7G.
|
| Description |
polydT-primed cDNA from total RNA
|
| Data processing |
Signal values for Affymetrix expression arrays were estimated using the GC-RMA module from Genedata’s Refiner 4.5 package (Genedata, Basel, Switzerland). Expression data analysis was performed in Genedata’s Analyst 4.5 package. Only those genes which had an Affymetrix detection p-value < 0.05 and signal values > 0.5 were assumed to be expressed.
|
| |
|
| Submission date |
Sep 14, 2009 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Michaela Schwaiger |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Vienna
|
| Lab |
Technau
|
| Street address |
Althanstr. 14
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL1322 |
| Series (1) |
| GSE18092 |
Heterochromatin protein 1 (HP1) modulates replication timing of Drosophila heterochromatin |
|
| Relations |
| Reanalyzed by |
GSE119084 |
