NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM2358944 Query DataSets for GSM2358944
Status Public on Jun 20, 2017
Title chip4_0Cu_Cy5_75Cu_Cy3 Replicate 2
Sample type RNA
 
Channel 1
Source name Seedlings
Organism Oryza sativa
Characteristics treatment: 0µM CuSO4
variety: Nippombare
age: 8 days
Treatment protocol Rice (Oryza sativa subsp japonica), cv. Nipponbare, seeds were surface-sterilized and stratified for 2 d at 4° C, and then germinated on ½MS plates with 1% Sucrose (Murashige and Skoog 1962). Two different Cu concentration conditions were compared, slight Cu deficiency (0 µM CuSO4) and Cu excess (75 µM CuSO4).
Growth protocol The plants were cultured in plates during 8 days under diferent concentration of CuSO4 in the medium.
Extracted molecule total RNA
Extraction protocol Total RNAs were isolated with the RNeasy plant mini kit (QIAgen, Hilden, Germany; Ref. 74904), following the manufacturer’s instructions. The aRNA amplification and labeling were performed with the AminoAllyl MessageAmpTM II aRNA Amplification kit (Ambion) according to the manufacturer’s instructions. One microgram of total RNA of each sample was used for the amplification. The final concentration of obtained aRNA was determined with NanoDrop ND1000 Spectrophotometer.
Label Cy5
Label protocol The obtained amplified RNA (aRNA) was labeled by using the CyDye Post-Labeling Reactive Dye Pack, which generates fluorescent Cy3- and Cy5-labeled probes by a post-labeling (amino allyl) method (GE Healthcare). Each biological replicate was labeled with Cy3 and Cy5 to produce four pairs of replicate dye-swaps, which were used for microarray hybridization
 
Channel 2
Source name Seedlings
Organism Oryza sativa
Characteristics treatment: 75µM CuSO4
variety: Nippombare
age: 8 days
Treatment protocol Rice (Oryza sativa subsp japonica), cv. Nipponbare, seeds were surface-sterilized and stratified for 2 d at 4° C, and then germinated on ½MS plates with 1% Sucrose (Murashige and Skoog 1962). Two different Cu concentration conditions were compared, slight Cu deficiency (0 µM CuSO4) and Cu excess (75 µM CuSO4).
Growth protocol The plants were cultured in plates during 8 days under diferent concentration of CuSO4 in the medium.
Extracted molecule total RNA
Extraction protocol Total RNAs were isolated with the RNeasy plant mini kit (QIAgen, Hilden, Germany; Ref. 74904), following the manufacturer’s instructions. The aRNA amplification and labeling were performed with the AminoAllyl MessageAmpTM II aRNA Amplification kit (Ambion) according to the manufacturer’s instructions. One microgram of total RNA of each sample was used for the amplification. The final concentration of obtained aRNA was determined with NanoDrop ND1000 Spectrophotometer.
Label Cy3
Label protocol The obtained amplified RNA (aRNA) was labeled by using the CyDye Post-Labeling Reactive Dye Pack, which generates fluorescent Cy3- and Cy5-labeled probes by a post-labeling (amino allyl) method (GE Healthcare). Each biological replicate was labeled with Cy3 and Cy5 to produce four pairs of replicate dye-swaps, which were used for microarray hybridization
 
 
Hybridization protocol From http://ag.arizona.edu/microarray/Microarraymethod1.doc Microarrays were re-hydrated by exposing them to water vapor for 10 seconds (four times). Next microarrays were cross-linked by ultraviolet (UV) irradiation of 180 mJ in a Stratalinker 1800 UV (Stratagene). Subsequently, microarrays were washed with 1% SDS (w/v) (Sodium Dodecyl Sulfate) for 5 min and 10 times with H2O milliQ (Millipore), washed with 100% ethanol 5 times and then dried by centrifugation at 200 g. After this pre-treatment, microarrays were hybridized overnight at 55 °C with hybridization solution [48 pmol of each labeled sample; deionized formamide 50% (v/v);] 3X SSC (Saline-Sodium Citrate); Denhardt´s solution [5 X and SDS 0.1% (w/v)]. The hybridization solution was denaturalized for 5 min at 65 °C and applied between the microarray and a coverslip LifterSlipTM (Erie Scientific).
Scan protocol The hybridized microarray slides were washed and scanned using a Gene Pix Autoloader (Axon/Molecular Devices, 4200A01, Sunnyvale, CA). Spot finding and data extraction was done using GenePix Pro 6 software (Axon/Molecular Devices, Sunnyvale, CA).
Data processing Quality control, normalization and determination of the differentially expressed genes were conducted in R using the Limma package (Smyth 2005), as previously described in (Andrés et al. 2009). To determine the false-positive ratio (FDR), the P values were adjusted by a test multiple test according to the method of Benjamini and Hochberg´s (Benjamini and Hochberg 1995). The genes that showed a differential expression with fold-change (FC) minimum of 2 for both conditions, represented as log2(FC) ≥ |1|of > 2-fold (-1 > log-fold change [½MS + 75 µM/ ½MS] > 1) with a P value of 0.05 (FDR = 0.5%) were selected.
 
Submission date Oct 24, 2016
Last update date Jun 20, 2017
Contact name Fernando Andres
E-mail(s) andres@mpipz.mpg.de
Organization name Max Planck Institute for Plant Breeding Research
Department Plant Developmental Biology
Lab Prof George Coupland
Street address Carl von Linne 10
City Cologne
ZIP/Postal code 50829
Country Germany
 
Platform ID GPL8756
Series (1)
GSE89109 Global tanscriptome analysis in Oryza sativa identifies genes regulated by copper in the medium

Data table header descriptions
ID_REF
VALUE Log2 fold-change (M-value) ratio (75 µM CuSO4/0 µM CuSO4)

Data table
ID_REF VALUE
TR030001 0.01648272
TR030002 0.178873132
TR030003 -1.095442704
TR030004 -0.157199563
TR030005 0.360826912
TR030006 -0.236894653
TR030007 -0.449500667
TR030008 0.112188354
TR030009 0.015775252
TR030010 0.273556422
TR030011 -0.910552113
TR030012 -0.33056734
TR030013 -0.085509192
TR030014 0.069510575
TR030015 0.4344078
TR030016 0.611094671
TR030017 0.138647103
TR030018 0.18059483
TR030019 0.400198089
TR030020 0.77182244

Total number of rows: 43311

Table truncated, full table size 904 Kbytes.




Supplementary file Size Download File type/resource
GSM2358944_chip4.gpr.gz 4.1 Mb (ftp)(http) GPR
Processed data included within Sample table

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