NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM184508 Query DataSets for GSM184508
Status Public on Dec 19, 2007
Title Pericycle root cells 2hr transitory KNO3 treated, biological rep3
Sample type RNA
 
Source name FACS-sorted 12 day old Arabidopsis Pericycle root cells, 2 hr KNO3 transitory treated
Organism Arabidopsis thaliana
Characteristics Genotype: Col-0
Cell type: pericycle root cells
Biomaterial provider Poethig Enhancer trap collection (http://enhancertraps.bio.upenn.edu/)
Treatment protocol At dawn on 12th day plants were either transitory/continuous nitrogen-treated (KNO3 to 5mM added) or KCl treated (KCl to 5mM added) as a control treatment for 2 hrs. To isolate single cell types (LRC, EpiC, EndoP, Peri, Stele) cells were incubated in a solution containing cell-wall digesting enzymes and then Fluorescence Activated Cell Sorting (FACS) sorted as Birnbaum et al (2005) Nature Methods 2: 615-619. To isolate mock FACS-sorted protoplasts the same proceedure was followed but the cells were not FACS-sorted, instead the protoplasts were incubated on ice for the time it takes to FACS-sort (1 hour). After FACS or mock-FACS cell sorting cells were frozen using liquid nitrogen. 5mM KNO3 was either maintained in the protoplast-generating solution (continuous treat) or not (transitory treat) to test how to best preserve the cellular nitrogen-response during the 1 hr protoplast generating procedure. As well as the 'protoplasts' controls we harvested whole roots that were either frozen using liquid nitrogen directly after the 2hr treatment or 3.5hr treatment, or whole roots incubated in protoplast-generating solultion (minus enzymes; transitory or continuous treated) and then frozen. 1mM MSX in combination with 5mM KNO3 (or with 5mM KCl as a control), or also with 5mM glutamine (Gln) for 2 hrs was used to block and then release the block of the assimilation of KNO3 in order to seperate KNO3 effects from assimilated-nitrogen effects in FACS-sorted pericycle cells.
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. All plant lines are in the Col-0 background.
Extracted molecule total RNA
Extraction protocol Total RNA was either extracted using the Plant RNAeasy kit (Qiagen, Hilden, Germany) for cell samples (LRC, EpiC, EndoP, Peri, Stele, Protoplasts), or for whole root samples using TRIzol reagent (Invitrogen, Carlsbad, CA). Protocols were followed according to manufacturer's instructions.
Label biotin
Label protocol Biotinylated cRNA from all isolated cell samples (LRC, EpiC, EndoP, Peri, Stele) were prepared according to the standard two-cycle Affymetrix protocol from 50 ng total RNA, or from all whole root samples and protoplast samples 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 protocol. 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 KNO3 was not maintained in the protoplast-generating solution
Data processing The data were analyzed with dChip using dChip default analysis settings and 'Average' as the normalization model, PM match only. Note that samples were normalized in two batches according to the sample type: Cell type specific samples (columns B-BC), and protoplasts and whole roots (columns BD-CG), all using the procedure described above. Non nuclear-encoded or ambiguous probe-gene matches were not used (according to latest annotation file from Affymetrix (April 2007). If the signal was log2 less than 6 in both treat and control experiments probes were removed from analysis; the value of 6 based on expression values of previously documented cell-specific genes. Probes determined to be affected by protoplast generation procedure were not analysed.
 
Submission date Apr 26, 2007
Last update date Aug 28, 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)
GSE7631 Cell-specific nitrogen responses in the Arabidopsis root
Relations
Reanalyzed by GSE118579
Reanalyzed by GSE119083

Data table header descriptions
ID_REF
VALUE dChip.2006 log2 normalized signal value

Data table
ID_REF VALUE
AFFX-BioB-5_at 12.739
AFFX-BioB-M_at 12.982
AFFX-BioB-3_at 12.801
AFFX-BioC-5_at 13.365
AFFX-BioC-3_at 13.153
AFFX-BioDn-5_at 13.396
AFFX-BioDn-3_at 13.775
AFFX-CreX-5_at 13.961
AFFX-CreX-3_at 14.014
AFFX-DapX-5_at 9.58
AFFX-DapX-M_at 12.739
AFFX-DapX-3_at 13.709
AFFX-LysX-5_at 8.53
AFFX-LysX-M_at 11.18
AFFX-LysX-3_at 13.129
AFFX-PheX-5_at 8.953
AFFX-PheX-M_at 11.161
AFFX-PheX-3_at 13.014
AFFX-ThrX-5_at 8.986
AFFX-ThrX-M_at 10.772

Total number of rows: 22810

Table truncated, full table size 357 Kbytes.




Supplementary file Size Download File type/resource
GSM184508.CEL.gz 2.1 Mb (ftp)(http) CEL
GSM184508.EXP.gz 282 b (ftp)(http) EXP
Raw data provided as supplementary file
Processed data included within Sample table

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap