Investigation Title	Transcription profiling of greenhouse grown wheat treated with three antifungal compounds (BTH, Azoxystrobin or Fenpropimorph)
Comment[Submitted Name]	Fungicide treatment
Experimental Design	compound_treatment_design	transcription profiling by array
Experimental Design Term Source REF	mo	EFO
Comment[ArrayExpressReleaseDate]	2005-06-01
Comment[AEMIAMESCORE]	4
Comment[ArrayExpressAccession]	E-MEXP-314
Comment[MAGETAB TimeStamp_Version]	2011-06-30 21:22:09 Last Changed Rev: 14857
Experimental Factor Name	time	compound
Experimental Factor Type	time	compound_treatment_design
Experimental Factor Term Source REF
Person Last Name	Zarn	Isidore	Keller	Pasquer
Person First Name	Jürg	Edwige	Beat	Frédérique
Person Mid Initials
Person Email				fpasquer@botinst.unizh.ch
Person Phone				+41 1 634 82 23
Person Fax
Person Address				Zollikestrasse 107, Zürich, Zürich, 8008, Switzerland
Person Affiliation				Plant Biology
Person Roles				submitter
Person Roles Term Source REF
Quality Control Type
Quality Control Term Source REF
Replicate Type
Replicate Term Source REF
Normalization Type
Normalization Term Source REF
Date of Experiment
Public Release Date	2005-06-01
PubMed ID	15774621
Publication DOI	15774621
Publication Author List	Jean Soulier, Emmanuelle Clappier, Jean-Michel Cayuela, Armelle Regnault, Marina García-Peydró, Hervé Dombret, André Baruchel,Maria-Luisa Toribio, and François Sigaux
Publication Title	HOXA genes are included in genetic and biological networks defining human acute T-cell leukemia (T-ALL)
Publication Status	journal_article
Publication Status Term Source REF	mo
Experiment Description	Effect of three antifungal compounds (BTH, Azoxystrobin or Fenpropimorph) on wheat gene expression.
Protocol Name	P-MEXP-7223	P-MEXP-8296	P-MEXP-8294	P-MEXP-8295	P-MEXP-7228	P-MEXP-8261	P-MEXP-8262	P-MEXP-7231	P-MEXP-8313
Protocol Type	grow	specified_biomaterial_action	specified_biomaterial_action	specified_biomaterial_action	nucleic_acid_extraction	labeling	labeling	hybridization	bioassay_data_transformation
Protocol Description	Wheat plants were grown in greenhouse (16hr light/20°C, 8hr night/16°C) till flag leaf stage.	Wheat plants were treated with fenpropimorph (Corbel, BASF, 1l/ha). Samples were collected 24hrs, one week and two weeks after treatments.	Wheat plants were treated with azoxystrobin (Amistar, Syngenta, 1l/ha). Samples were collected 24hrs, one week and two weeks after treatments.	Wheat plants were treated with BTH (Bion, Novartis, 60g/ha). Samples were collected 24hrs, one week and two weeks after treatments.	Total RNA of the samples was extracted using the TRizol method. 1g of leaf material was ground in liquid nitrogen and 10 ml of TRizol (38% phenol, guanidine thiocyanate 0.8 M, ammonium thiocyanate 0.4 M, sodium acetate pH 5 0.1 M, glycerol 5%) was added, mixed thoroughly, and incubated for 5 min at room temperature (RT). After centrifugation for 15 min at 12,000 g (4°C), the supernatant was transferred into a new tube and 2 ml of chloroform were added and mixed 15 sec using a vortex. After 2 min incubation at RT, samples were centrifuged for 15 min at 12,000 g (4°C). The supernatant (aqueous phase) was pipetted to a new tube. 0.5 volume of isopropanol and 0.5 volume of 0.8 M sodium citrate/1.2 M NaCl were added and mixed by inversion. After 1hr incubation at -20°C, samples were precipitated by centrifugation (10 min at 10,000 g at 4°C). Pellets were washed in 10 ml 75% ethanol, centrifuged for 10 min at 10,000 g (4°C), dried and resuspended in 300 ml of DEPC-treated ddH2O. Samples were subsequently submitted to another purification step using a QIAshredder column (Qiagen) 	Total RNA from control plants was directly used as template for reverse transcription using a poly-dT primer of 21 nucleotides. 40 ug of total RNA were denatured at 70°C for 5 min together with 2 ug of primer in a total volume of 13 ul. After incubation for 5 min at RT, the reaction mix containing 2 ul of Superscript II, 6 ul of 5x reaction buffer (Invitrogen Life Technologies, Basel, Switzerland), 3 ul of 0.1 M DTT, 0.8 ul of RNase Out 40 U/ul (Invitrogen Life Technologies, Basel, Switzerland), 0.6 ul of 25 mM dATP, dGTP and dTTP, 0.6 ul of 10 mM dCTP and 3 ul of Cyanine3-dCTP (Cy3,Amersham, Otelfingen, Switzerland) was added and incubated for 2 hrs at 42°C. 	Total RNA from treated plants was directly used as template for reverse transcription using a poly-dT primer of 21 nucleotides. 40 ug of total RNA were denatured at 70°C for 5 min together with 2 ug of primer in a total volume of 13 ul. After incubation for 5 min at RT, the reaction mix containing 2 ul of Superscript II, 6 ul of 5x reaction buffer (Invitrogen Life Technologies, Basel, Switzerland), 3 ul of 0.1 M DTT, 0.8 ul of RNase Out 40 U/ul (Invitrogen Life Technologies, Basel, Switzerland), 0.6 ul of 25 mM dATP, dGTP and dTTP, 0.6 ul of 10 mM dCTP and 3 ul of Cyanine5-dCTP (Cy5, Amersham, Otelfingen, Switzerland) was added and incubated for 2 hrs at 42°C. 	Before hybridisation, 1.25 ul of 10 ug/ul yeast RNA, 1.9 ul 20x SSC and 0.5 ul of 10% SDS were added to the labelled cDNA. Labelled samples were added to the microarray which was covered with a cover-slip and placed into a pre-warmed hybridisation chamber containing 20 ul of 3x SSC in its wells.	Spots flagged as empty by the software Imagene 4.2 or manually were removed from the analysis. Normalisation of the signal intensities between the two channels was performed using the global method, and between the slides by scale normalisation as described in Yang and Dudoit, 2002.
Protocol Parameters					Extracted product;Amplification;	Amount of nucleic acid labeled;Label used;Amplification;	Amplification;Label used;Amount of nucleic acid labeled;	temperature;time;Volume;Chamber type;Quantity of label target used;
Protocol Hardware
Protocol Software
Protocol Contact
Protocol Term Source REF					mo	mo	mo	mo	mo
SDRF File	E-MEXP-314.sdrf.txt
Term Source Name	gramene		ArrayExpress	mo	EFO
Term Source File	http://www.gramene.org/plant_ontology/index.html		http://www.ebi.ac.uk/arrayexpress	http://mged.sourceforge.net/ontologies/MGEDontology.php	http://www.ebi.ac.uk/efo/
Term Source Version