|
| Status |
Public on Sep 01, 2013 |
| Title |
Ts5 roots 2hr KCl control treated, biological rep3 |
| Sample type |
RNA |
| |
|
| Source name |
12 day old Arabidopsis Ts5 ecotype roots, 2 hr 5mM KCl control treated
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
tissue: root
ecotype: Ts5
agent: KCl control
age: 12 day old seedling roots
|
| Treatment protocol |
At dawn on 12th day plants were either nitrogen-treated (KNO3 to 5mM added) or KCl treated (KCl to 5mM added) as a control treatment for 2 hrs. Whole roots were then harvested and frozen using liquid nitrogen.
|
| Growth protocol |
Plants were grown for 12 days in long day (16hr light/8hr dark) 22 oC conditions in a Percival (Percival Scientific). Plants were grown hydroponically in basal MS containing no nitrogen except 0.5mM ammonium succinate and 3mM sucrose.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using TRIzol reagent (Invitrogen, Carlsbad, CA) according to manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA from all isolated root samples were prepared according to the standard one-cycle Affymetrix protocol from 1 ug total RNA.
|
| |
|
| Hybridization protocol |
Following fragmentation, 8 ug of cRNA were hybridized for 16 hr at 45C on an Affymetrix ATH1 GeneChip Arabidopsis Whole Genome Array in the Affymetrix Hybridization oven 640 following standard Affymetrix protocols. GeneChips were washed and stained using the 'EukGE-WS2v4_450' 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 |
3 Ts5 C.CEL
|
| Data processing |
The data were analyzed with MAS5.0 as normal for Col0 data but with a modification to the normalisation method to enable a subset of probe sequences within each probe set to be excluded for non-Col0 data. Probe sequences (within probe sets for each gene) whose signal values were not used were those predicted using our bioinformatic analysis not to hybridise non-Col0 RNA on the Col0 Affymetrix microarray due to sequence differences in the non-Col0 ecotypes; these were excluded to give a more accurate signal value for each gene in the non-Col0 ecotypes. The probe sequences to be excluded were identified per experiment (i.e. an experiment consisted of the three replicate samples, e.g. Kas2 control, or Kas2 treat). Lists of probe sequences within each gene probeset that were not used for normalisation are supplied in text files (one per experiment) together with a key that denotes the probe numbers given in the experiment files to the Affymetrix probe IDs. On average, 10% of all probe sets were analysed with the complete set of 11 elements and a further 70% analysed with 9 or 10 elements. For example, in Kas2C_Probe_Elements_Removed, the first row is 4,1. 4,1 refers to probe number '4'. The key for which probe number '4' is, is in the 'key' file and it corresponds to the probe name given on row 4. The same key file is used to interpret all of the other files. The '1' means that the first element of this probeset was excluded from analysis. The data was analysed within replicate sets, so for replicates 1, 2, and 3 of Kas2 C the same elements were removed. The reasoning for this is that we used all replicates to identify with best accuracy which elements were not hybridising normally.
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| |
|
| Submission date |
Dec 04, 2011 |
| Last update date |
Aug 15, 2018 |
| Contact name |
Kenneth David Birnbaum |
| E-mail(s) |
ken.birnbaum@nyu.edu
|
| 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) |
| GSE34130 |
Ecotype specific nitrogen responses in the Arabidopsis root |
|
| Relations |
| Reanalyzed by |
GSE118579 |
