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Sample GSM142594 Query DataSets for GSM142594
Status Public on Jan 22, 2007
Title DB001_ATH1_A4-Brown-cal
Sample type RNA
 
Source name A-4-Brown-9day_high
Organism Arabidopsis thaliana
Characteristics OrganismPart: Cell culture
Age time point: 9 day timepoint
Ecotype: Ler
NASC:StockCode: NW20
Treatment protocol Leaves from seedlings, grown aseptically (14-17 days)on MS agar, excised and transferred to callus induction medium (Gamborg's B5 Basil Medium with minimal organics, 2.0% glucose, 0.5mgl-1 MES, 0.5mgl-1 2,4-D, 0.05mgl-1 Kinetin. pH 5.7). Incubated in darkness, at 22oC for 2 weeks. Callus transferred to high differentiating liquid media (MS 0.2% sucrose, 4% mannitol, 0.2mgl-1 auxins (2,4-D, NAA, IAA) 1mgl-1 BA, pH5.7). Incubated at 22oC on a rotary shaker (105rpm). Sample taken at day =9 ; Performer: David,,Brown
Growth protocol unknown; Parameter location = Growth Room; Parameter Temperature = 22oC; Parameter Medium = Murahige and Skoog; Parameter Treatment = 0.2mgl-1 auxin( 2,4-D NA IAA) 1.0mgl-1 BA; Parameter Nutrients = 0.2% sucrose 4% mannitol; Performer: David,,Brown
Extracted molecule total RNA
Extraction protocol Qiagen Plant RNAeasy kit; Protocol: Seedlings harvested and snap-frozen in liquid nitrogen and RNA extracted as described in the Qiagen RNAeasy kit. ; Protocol Type = nucleic_acid_extraction; Performer: David,,Brown
Label biotin
Label protocol Preparation of biotin-labeled cRNA; RNA samples were quality controlled using the Agilent 2100 Bioanalyzer. 100 pmol T7-(dT)24 primer (5'-GGCCAGTGAATTGTAATACGACTCACTATAGGGAGGCGG-(dT)24-3') was hybridized with the Total RNA ( 8 - 12ug) at 70°C for 10 minutes. First strand cDNA was synthesized by reverse transcription using 400 units SuperScript II Reverse Transcriptase in a reaction containing 50 mM Tris-HCl (pH 8.3), 75 mM KCl, 3 mM MgCl2, 10 mM dithiothreitol (DTT) and 0.5 mM dNTPs at 42C for 1 hour. The reaction was then put on ice for 2 mins, followed by second strand cDNA synthesis. 130ul of the second strand reaction mix was added to the 20ul first strand synthesis reaction. The second strand reaction mix contained 40 units E. coli DNA polymerase I, 10 units E. coli DNA ligase, 2 units E. coli RNase H and 0.2 mM dNTPs in a buffer containing 20 mMTris-HCl (pH 6.9), 90 mM KCl, 4.6 mM MgCl2, 10mM (NH4)SO4 and 0.15 mM ß-NAD+. The reaction was incubated at 16 C for 2 hours. 10 units T4 DNA Polymerase was added and the reaction incubated for a further 5 minutes at 16 C, then the reaction was terminated using EDTA. The double-stranded cDNA was cleaned up using the cDNA Cleanup Spin Column supplied in the Affymetrix GeneChip Sample Cleanup Module. The double-stranded cDNA was used as a template for in Vitro transcription of biotin-labeled cRNA using the ENZO BioArray RNA Transcript Labeling Kit supplied by Affymetrix. The Transcript Labeling kit produces large amounts of hybridizable biotin-labeled RNA targets by in Vitro transcription from bacteriophage T7 RNA polymerase promoters. The reaction mixture containing Biotin-Labeled ribonucleotides (ATP, GTP, CTP, UTP with Bio-UTP and Bio-CTP), DTT, RNase inhibitor mix and T7 RNA polymerase was incubated at 37 C for 5 hrs, gently mixing every 30 minutes during the incubation. This produced an average of 40ug biotin-labeled cRNA . The biotin-labeled cRNA was cleaned up using the cRNA Cleanup Spin Column supplied in the Affymetrix GeneChip Sample Cleanup Module. The quantity and quality of the cRNA was measured using the Agilent 2100 Bioanalyzer. The total amount of cRNA was calculated, the starting quantity of RNA subtracted to give the quantity of adjusted cRNA. The cRNA was fragmented by metal-induced hydrolysis to break down full-length cRNA to 35-200 base fragments. 15ug of adjusted cRNA was used to prepare 300ul of hybridization cocktail of which 200ul was hybridized with the GeneChip. ; Protocol Type = labeling; Performers: John,Y,Okyere, Janet,,Higgins
 
Hybridization protocol EukGE-WS2v4; Protocol Type = Hybridization; Parameter Wash A1 Recovery Mixes = 0 (type integer); Parameter Wash A1 Temperature (C) = 25 (type integer); Parameter Number of Wash A1 Cycles = 10 (type integer); Parameter Mixes per Wash A1 Cycle = 2 (type integer); Parameter Wash B Recovery Mixes = 0 (type integer); Parameter Wash B Temperature (C) = 50 (type integer); Parameter Number of Wash B Cycles = 4 (type integer); Parameter Mixes per Wash B Cycle = 15 (type integer); Parameter Stain Temperature (C) = 25 (type integer); Parameter First Stain Time (seconds) = 600 (type integer); Parameter Wash A2 Recovery Mixes = 0 (type integer); Parameter Wash A2 Temperature (C) = 25 (type integer); Parameter Number of Wash A2 Cycles = 10 (type integer); Parameter Mixes per Wash A2 Cycle = 4 (type integer); Parameter Second Stain Time (seconds) = 600 (type integer); Parameter Third Stain Time (seconds) = 600 (type integer); Parameter Wash A3 Recovery Mixes = 0 (type integer); Parameter Wash A3 Temperature (C) = 30 (type integer); Parameter Number of Wash A3 Cycles = 15 (type integer); Parameter Mixes per Wash A3 Cycle = 4 (type integer); Parameter Holding Temperature (C) = 25 (type integer); Software: type: Acquisition and analysis software; Hardware: type: Fluidics Station, Parameter: Parameter Station Number = 1 (type integer), Parameter: Parameter Position = 2 (type integer); Performers: John,Y,Okyere, , Janet,,Higgins
Scan protocol Protocol Type = CEL Analysis; Parameter Algorithm name = Percentile (type string); Parameter Upper left corner x coordinate = 230 (type integer); Parameter Upper left corner y coordinate = 232 (type integer); Parameter Upper right corner x coordinate = 4498 (type integer); Parameter Upper right corner y coordinate = 259 (type integer); Parameter Lower right corner x coordinate = 4472 (type integer); Parameter Lower right corner y coordinate = 4530 (type integer); Parameter Lower left corner x coordinate = 203 (type integer); Parameter Lower left corner y coordinate = 4502 (type integer); Parameter Percentile = 75 (type integer); Parameter Cell margin = 2 (type integer); Parameter Outlier high = 1.500 (type float); Parameter Outlier low = 1.004 (type float); Software: type: Acquisition and analysis software;
Protocol Type = Scan; Parameter Pixel Size = 3 (type integer); Parameter Number of Scans = 2 (type integer); Software: type: Acquisition and analysis software; Hardware: type: Scanner, Parameter: Parameter Scanner ID = not specified (type string), Parameter: Parameter Scanner Type = HP (type string); Performers: John,Y,Okyere, , Janet,,Higgins
Description Rows (integer) = 712
Cols (integer) = 712
Number of Probe Sets (integer) = 22810
Background Avg (float) = 74.33
Background Stdev (float) = 1.92
Background Max (float) = 78.4
Background Min (float) = 69.0
Noise Avg (float) = 4.43
Noise Stdev (float) = 0.14
Noise Max (float) = 5.0
Noise Min (float) = 4.0
RawQ (float) = 2.84
Number Cells Masked (integer) = 0
Number Outlier Cells (integer) = 254
Number Cells Modified (integer) = 0
Rows (integer) = 712
Cols (integer) = 712
Number of Cells (integer) = 506944
Image: DB001_ATH1_A4-Brown-cal.DAT
Data processing Protocol Type = CHP Analysis; Parameter Algorithm name = ExpressionStat (type string); Parameter Algorithm version = 5.0 (type string); Parameter Baseline file = not specified (type string); Parameter Alpha1 = 0.04 (type float); Parameter Alpha2 = 0.06 (type float); Parameter Tau = 0.015 (type float); Parameter Gamma1H = 0.0025 (type float); Parameter Gamma1L = 0.0025 (type float); Parameter Gamma2H = 0.003 (type float); Parameter Gamma2L = 0.003 (type float); Parameter Perturbation = 1.1 (type float); Parameter Normalization factor = 1.000000 (type float); Parameter Scale factor = 0.377090 (type float); Parameter Mask file = not specified (type string); Software: type: Acquisition and analysis software;
 
Submission date Oct 27, 2006
Last update date Aug 28, 2018
Contact name Nottingham Arabidopsis Stock Centre (NASC)
E-mail(s) affy@arabidopsis.info
Phone +44 (0)115 951 3237
Fax +44 (0)115 951 3297
URL http://arabidopsis.info/
Organization name Nottingham Arabidopsis Stock Centre (NASC)
Department School of Biosciences, University of Nottingham
Street address Sutton Bonington Campus
City Loughborough
ZIP/Postal code LE12 5RD
Country United Kingdom
 
Platform ID GPL198
Series (1)
GSE6148 The trans-differentiation of cultured Arabidopsis cells
Relations
Reanalyzed by GSE119083

Data table header descriptions
ID_REF Type: string; Scale: OTHER
VALUE MAS5.0
ABS_CALL
DETECTION P-VALUE

Data table
ID_REF VALUE ABS_CALL DETECTION P-VALUE
AFFX-BioB-5_at 45.6 P 0.02611
AFFX-BioB-M_at 56.7 P 0.00006
AFFX-BioB-3_at 32.7 P 0.00141
AFFX-BioC-5_at 103.4 P 0.00030
AFFX-BioC-3_at 54.9 P 0.00034
AFFX-BioDn-5_at 99.8 P 0.00004
AFFX-BioDn-3_at 482.8 P 0.00006
AFFX-CreX-5_at 1378.8 P 0.00004
AFFX-CreX-3_at 1582.7 P 0.00004
AFFX-DapX-5_at 3.8 A 0.38260
AFFX-DapX-M_at 6.3 A 0.41138
AFFX-DapX-3_at 0.7 A 0.92732
AFFX-LysX-5_at 0.6 A 0.64555
AFFX-LysX-M_at 1.6 A 0.86864
AFFX-LysX-3_at 4.6 A 0.22764
AFFX-PheX-5_at 0.9 A 0.92730
AFFX-PheX-M_at 1.1 A 0.83414
AFFX-PheX-3_at 2.1 A 0.94156
AFFX-ThrX-5_at 0.7 A 0.86052
AFFX-ThrX-M_at 2.9 A 0.57404

Total number of rows: 22810

Table truncated, full table size 561 Kbytes.




Supplementary file Size Download File type/resource
GSM142594.CEL.gz 2.3 Mb (ftp)(http) CEL
Raw data provided as supplementary file

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