|
| Status |
Public on Mar 01, 2009 |
| Title |
IAA7mII over-expressor_mock treated_replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
IAA7mII over-expressor, mock treated
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
Genotype: Col-0
Pwer:GFP positive protoplasts
|
| Biomaterial provider |
Courtesy of John Schiefelbein
|
| Treatment protocol |
The roots of the seedlings were harvested with a scalpel and incubated in protoplasting solution for 3 h. Protoplasts were filtered and PEG-transformed with an empty vector (pMON999_mRFP), an IAA7mII over-expressor (pBeaconRFP_IAA7mII) or an IAA19mII over-expressor (pBeaconRFP_IAA19mII). Protoplasts were left overnight and treated for 3 h with 5microM IAA or solvent alone. Dually labeled protoplasts were sorted directly into RNA extraction buffer by FACS.
|
| Growth protocol |
Pwer::GFP plants were grown in vertically placed square 10x10 cm petri dishes for 7 days in long day (18hr light/6hr dark) 22C conditions in a Percival (Percival Scientific). Plants were grown on MS agar (0.5x MS, 1% sucrose, 1% agar, pH 5.7 with KOH) on top of a nylon mesh.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Protoplasts were sorted into 350 microliters RLT exrtraction buffer from a QIAGEN Rneasy micro kit and RNA was extracted according to the manufacturer's instructions, including a DNase treatment.
|
| Label |
biotin
|
| Label protocol |
cDNA was produced, amplified, fragmented and labeled using NuGEN WT-Ovation Pico RNA Amplification System and FL-Ovation cDNA Biotin Module V2.
|
| |
|
| Hybridization protocol |
Labeled cDNA was hybridized for 16 hr at 45C on an Affymetrix ATH1 GeneChip Arabidopsis Whole Genome Array in the Affymetrix Hybridization oven 640 following standard Affymetrix protocol. GeneChips were washed and stained using an Affymetrix Wash and Stain Kit and the FS450_001 protocol in the Affymetrix GeneChip Fluidics Station 450.
|
| Scan protocol |
GeneChips were scanned using the Affymetrix GeneChip 3000 Scanner according to standard Affymetrix protocols.
|
| Description |
Roots grown horizontally on square plates, protoplasted, transformed and sorted by FACS.
Replicate 3 of 3.
|
| Data processing |
The data was normalized using MAS5 with a target intensity of 250.
|
| |
|
| Submission date |
Dec 01, 2008 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Kenneth David Birnbaum |
| E-mail(s) |
[email protected]
|
| Phone |
212-998-8257
|
| Organization name |
New York University
|
| Department |
Biology
|
| Lab |
Birnbaum
|
| Street address |
12 Waverly Place
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10003 |
| Country |
USA |
| |
|
| Platform ID |
GPL198 |
| Series (1) |
| GSE13783 |
Positive fluorescent selection permits precise, rapid and in-depth over-expression analysis in plant protoplasts |
|
| Relations |
| Reanalyzed by |
GSE118579 |
| Reanalyzed by |
GSE119083 |
