|
| Status |
Public on Aug 24, 2018 |
| Title |
wt1:N+.1 |
| Sample type |
RNA |
| |
|
| Source name |
Strain wt1, 60mM Gln, replicate 1
|
| Organism |
Fusarium fujikuroi |
| Characteristics |
treatment: 60mM Gln
|
| Treatment protocol |
The mycelium was harvested, shock frozen with liquid nitrogen and lyophilized.
|
| Growth protocol |
Strains were grown for 3 days in Darken medium and subsequently, an aliquot of 500 µL was transferred to liquid ICI medium with either 6 mM or 60 mM glutamine and grown for additional 3 days on a rotary shaker (180 rpm) at 28 °C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the TRI Reagent (Sigma-Aldrich, Steinheim, Germany) and DNA was removed using DNAse I (Roche Diagnostics, Mannheim, Germany). RNA quality and concentration was measured with an Bioanalyzer (Agilent Technologies Deutschland, Böblingen, Germany).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.5 ug of Cy3-labeled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60 °C for 30 minutes in a reaction volume of 250 mL containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturer's instructions. Upon completion of the fragmentation reaction, 250 mL of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65 °C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37 °C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10 um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Data processing |
Preprocessing steps including background correction, normalization and summarization of probe intensities were performed by employing the oligo R package (Carvalho 2010).
|
| |
|
| Submission date |
Dec 06, 2016 |
| Last update date |
Aug 24, 2018 |
| Contact name |
Ulrich Guldener |
| E-mail(s) |
u.gueldener@tum.de
|
| Organization name |
Technische Universität München
|
| Department |
Chair of genome-oriented bioinformatics
|
| Street address |
Maximus-von-Imhof-Forum 3
|
| City |
Freising |
| State/province |
Bavaria |
| ZIP/Postal code |
D-85354 |
| Country |
Germany |
| |
|
| Platform ID |
GPL22756 |
| Series (1) |
| GSE90948 |
Genome-wide transcriptome analysis of Fusarium fujikuroi Δset1 and Δkdm5 mutants |
|
