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Sample GSM147701 Query DataSets for GSM147701
Status Public on Nov 29, 2007
Title Vitis vinifera 'Cabernet sauvignon' mock inoculated at 48 h post inoculation 30
Sample type RNA
 
Source name The 10 randomly collected V. vinifera 'Cabernet Sauvignon' leaves at 48 hour after mock inoculation
Organism Vitis vinifera
Characteristics Fully-expanded leaves at the third or fourth position from the top of each shoot were used for treatment
Treatment protocol third or forth leaves were mock inoculated
Growth protocol 14 hours day/10 hours night. Light intensity: 500 umol m-2. Light source: high pressure sodium and metal halide. Temperature: 25°C. Relative humidity: 85%.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from each replicate sample individually. The leaves were ground in liquid nitrogen and homogenized in extraction buffer (2% Hexadecyltrimethyl Ammonium Bromide 1% sodium dodecyl sulfate, 2.5M NaCl, 0.5M Tris, 50mM EDTA, 5% beta-mercaptoethanol, and 3% polyvinyl poly-pyrrolidone. Following a 30-min incubation at -80°C, the samples were thawed and centrifuged. The supernatant was supplemented with 1/30 volume of 3 M sodium acetate (pH5.2) and 1/10 volume of ethanol, incubated on ice and centrifuged three times at 7000g, 30 min 4 ºC, each time transferring the supernatant into a clean tube. The supernatant was supplemented with 1/9 volume of 3 M sodium acetate (pH5.2) and iso-propanol to a final concentration of 33%. Following 60-min incubation at -20°C, the RNA was collected by centrifugation at 12,000g, 30 min, 4 ºC. The RNA was then supplemented with 1/3 volume of 8 M LiCl and incubated overnight at 4 °C. RNA was then collected by centrifugation at 12,000g, 30 min, 4 ºC, washed with 75% ethanol, and resuspended in 40 ul DEPC water. Ten micrograms of total RNA were treated with Turbo DNase I (Ambion) and purified with RNA purification kit (Qiagen) according to manufacturer’s instructions.
Label biotin
Label protocol The cDNA synthesis and antisense cRNA amplification/biotin labeling was performed using the One-Cycle Target Labeling Kit (Catalog number: 900493, Affymetrix, Santa Clara, California)
 
Hybridization protocol cRNA was cleaned up by the Affymetrix sample cleanup module which in included in the One-Cycle Target Labeling kit. Fifteen ug of cRNA at concentration of 1.301ug/ul were used for hybridization. Prior to hybridization, cRNA was heated at 94ºC for 35 min, and was fragmented to 35 to 200 nucleotides. After adding hybridization buffer to the fragmented samples, hybridization cocktails were boiled for 5 min, incubated for 5 min at 45ºC, then centrifuged for 5 minutes prior to loading per standard one-cycle protocol. Hybridization took place in an Affy hybridization oven model 640, for 16h at 45ºC. All washing took place on the fluidics station (as per the recommended fluidics protocol) with non-stringent wash buffer (6X SSPE and 0.01% Tween-20) and Stringent wash buffer (100mM MES, 0.1M NaCl, 0.01% Tween-20). Fluorescence was amplified by first adding streptavidin-phycoerythrin (SAPE) stain, then adding a biotinylated antibody (anti-streptavidin) solution, followed by another SAPE staining (as per standard one-cycle protocol). The fluidics protocol Midi_Euk2v3 was used (as per V. vinifera microarray package insert) on fluidics station model 450.
Scan protocol Affymetrix Scanner 3000 7G was used to collect at resolution of 1.56um with emission filter at a wavelength of 570nm and excitation wavelength of 532nm.
Description Gene expression data from V. vinifera 'Cabernet Sauvignon' at 0 hour post mock inoculation
Data processing Raw intensity data were processed using the GeneChip Operating Software Version 1.2 (GCOS 1.2; Affymetrix, 2001). Background correction and expression value calculation were performed as described by Affymetrix (2002). Normalization was done by global scaling, with a target intensity value of 150 (Affymetrix, 2001). Probe sets with missing values were deleted across all replicates and species to allow comparisons among datasets
 
Submission date Nov 30, 2006
Last update date Dec 06, 2006
Contact name Wenping Qiu
E-mail(s) wenpingqiu@missouristate.edu
Phone (417)5477517
Fax (417)5477540
URL http://mtngrv.missouristate.edu/CGB/
Organization name Missouri State University
Department Agriculture
Street address 9740 Red Spring Road
City Mountain Grove
State/province MO
ZIP/Postal code 65711
Country USA
 
Platform ID GPL1320
Series (1)
GSE6404 V. vinifera 'Cabernet sauvignon' and V. aestivalis 'Norton' innoculated with Erysiphe necator conidiospores

Data table header descriptions
ID_REF
VALUE GCOS-calculated Signal intensity
ABS_CALL the call in an absolute analysis that indicates if the transcript was present (P), absent (A), marginal (M), or no call (NC
DETECTION P-VALUE 'detection p-value', p-value that indicates the significance level of the detection call

Data table
ID_REF VALUE ABS_CALL DETECTION P-VALUE
AFFX-BioB-5_at 54.7 P 0.000127
AFFX-BioB-M_at 65.3 P 0.000052
AFFX-BioB-3_at 71.2 P 0.000044
AFFX-BioC-5_at 185.5 P 0.000052
AFFX-BioC-3_at 125.1 P 0.000052
AFFX-BioDn-5_at 382.2 P 0.000044
AFFX-BioDn-3_at 730 P 0.000044
AFFX-CreX-5_at 2706.6 P 0.000052
AFFX-CreX-3_at 3755.8 P 0.000052
AFFX-DapX-5_at 111.7 P 0.000044
AFFX-DapX-M_at 247.7 P 0.000052
AFFX-DapX-3_at 531.9 P 0.000044
AFFX-LysX-5_at 23 P 0.000052
AFFX-LysX-M_at 39.4 P 0.000127
AFFX-LysX-3_at 46 P 0.00007
AFFX-PheX-5_at 26.3 P 0.000169
AFFX-PheX-M_at 31.2 P 0.000258
AFFX-PheX-3_at 74.9 P 0.000052
AFFX-ThrX-5_at 36.9 P 0.00007
AFFX-ThrX-M_at 57.5 P 0.000044

Total number of rows: 16602

Table truncated, full table size 439 Kbytes.




Supplementary file Size Download File type/resource
GSM147701.cel.gz 1.9 Mb (ftp)(http) CEL
Raw data provided as supplementary file

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