Comment[ArrayExpressAccession] E-GEOD-51353 MAGE-TAB Version 1.1 Public Release Date 2013-10-18 Investigation Title The role of the Hog1 and Slt2 MAP Kinases in the regulation of Saccharomyces cerevisiae gene expression upon stress by sulfuric acid Comment[Submitted Name] The role of the Hog1 and Slt2 MAP Kinases in the regulation of Saccharomyces cerevisiae gene expression upon stress by sulfuric acid Experiment Description In our previous work, we had found that Saccharomyces cerevisiae needs of the Hog1 and Slt2 proteins to growth in a low pH environment caused by sulfuric acid, one of the stress factors during the process of ethanol production. Then was performed the gene-wide expression analysis in the hog1M-bM-^HM-^F and slt2M-bM-^HM-^F mutants in order to reveal the function of the Hog1p and Slt2p MAP Kinases in the regulation of S. cerevisiae global gene expression upon stress by sulfuric acid. BY4741 strain (Reference) and their derivate mutants hog1M-bM-^HM-^F and slt2M-bM-^HM-^F were grown for 1 h in YPD medium pH 2.5 adjusted with concentrated sulfuric acid Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Lucena Rodrigo Carolina Will Sergio Diogo Marcos Person First Name Rodrigo Lucena Elsztein Pita Paiva SimM-CM-5es Morais Person Mid Initials MendonM-CM-'a Person Email lucenarm2@gmail.com Person Affiliation Federal University of Pernambuco Person Address Genetics, Federal University of Pernambuco, Av. Jorn. AnM-CM--bal Fernandes, Recife, Pernambuco, Brazil Person Roles submitter Protocol Name P-GSE51353-1 P-GSE51353-5 P-GSE51353-6 P-GSE51353-2 P-GSE51353-3 P-GSE51353-4 P-GSE51353-7 Protocol Description Data were then analyzed using specific packages from the R statistical language [20] with the specific LIMMA and MArray packages. Background correction was done with normexp method. Robust spline and quantile methods were used for normalization within and between arrays, respectively. A list of differentially expressed genes was generated by applying adjusted p-values for multiple tests [20]. Groups of genes with statistical significance (Adj.p<0.05) were utilized. ID_REF = VALUE = normalized log10 ratio (Cy5/Cy3) representing test/reference Synthesis of cDNA and labelling was performed by Two-Color Low Input Quick Amp Labeling Kit (cat. nr. 5190-2306, Agilent, USA), with Two-Color RNA spike-in kit for internal control (cat. nr. 5188-5279, Agilent, USA). For yeast samples, 100 ng of purified RNA was used for labelling. RNA from lab strain BY4741 was labelled with Cy3 and mixed with Cy5-labelled RNA from hog1 and slt2 mutants. Target and reference cRNA samples were mixed and the hybridizations were performed on yeast gene expression 8x15K spots slides (cat. nr. G4813A-016322, Agilent, USA) at 65 M-:C for 17 hours at 10 r.pm. in the Microarray Hybridization Oven (cat. nr. G2545A, Agilent, USA). The cultures were centrifuged and each one was re-suspended with YPD medium pH 2.5 adjusted with sulfuric acid. After incubation for 1 h at 30M-0C with orbital agitation (150 rpm), the cells were recovered by centrifugation and immediately used to RNA extraction Cell from BY4741 strain (EUROSCARF collection) and their derivate mutants hog1M-bM-^HM-^F and slt2M-bM-^HM-^F were growth overnight and were transferred to fresh neutral YPD medium (20 g/l glucose, 20 g/l yeast extract and 10 g/l peptone) at 30M-:C and 150 rpm in a rotatory shaker at an initial cell density of OD600 0.1 and cultivated to OD600 0.5. total RNA in the lysates was purified by using RNAspin Mini RNA Isolation Kit (cat. nr. 25-0500-71, GE HealthCare, USA) following manufacturerM-4s instructions and suspended in 25 M-NM-