The prototrophic S. cerevisiae strain CEN.PK113-7D [18] was used for this study
Biomaterial provider
J Aguilera
Treatment protocol
Liquid nitrogen quenching
Growth protocol
Cells were grown at 30 °C in laboratory fermenters (Applikon, Schiedam, The Netherlands) with a working volume of 1 l as described in . Cultures were fed with a defined synthetic medium that was designed to allow for steady-state growth limited by either carbon or nitrogen, with all other requirements in excess and at a constant residual concentration. The dilution rate was set to 0.10 h−1. The pH was measured online and kept constant at 5.0 by the automatic addition of 2-M KOH with the use of an Applikon ADI 1030 Biocontroler. Stirrer speed was 800 rpm, and the gas flow was 0.5 l min−1. Synthetic media were prepared as described by Verdyun (1992) with the following modifications: for carbon-limited cultivation, the medium contained 5.0 g l−1 of (NH4)2SO4, 3.0 g l−1 of KH2PO4, 0.5 g l−1 of MgSO4 · 7H2O, and either 7.5 g l−1 of glucose or 5.76 g l−1 of ethanol. For nitrogen-limited cultures, 1.0 g l−1 of (NH4)2SO4, 5,3 g l−1 of K2SO4, 3.0 g l−1 of KH2PO4, 0.5 g l−1 of MgSO4 · 7H2O, and the necessary glucose to keep the residual glucose concentration at 18 g l−1 (59 and 62.2 g l−1 for the CO2-untreated and -treated cultures, respectively). This was done to avoid differences in the degree of glucose repression. For CO2-enriched aerobic cultivation, cultures were sparged with a defined gas mixture containing 79% CO2 and 21% O2 (HoekLoos, Schiedam, The Netherlands).
Extracted molecule
total RNA
Extraction protocol
One hundred millilitres of culture was sampled directly from the fermentor into a beaker containing 200–300 ml of liquid nitrogen and then processed . Total RNA was extracted with phenol/chloroform. mRNA extraction, cDNA synthesis and labelling, as well as array hybridisation against Affymetrix YG-S98 GeneChips® was performed as described in the Affymetrix user’s manual. Data acquisition was performed using the software packages Microarray Suite v5.0, MicroDB v3.0 and Data Mining Tool v3.0 (Affymetrix, Santa Clara, CA, USA).
One hundred millilitres of culture was sampled directly from the fermentor into a beaker containing 200–300 ml of liquid nitrogen and then processed . Total RNA was extracted with phenol/chloroform. mRNA extraction, cDNA synthesis and labelling, as well as array hybridisation against Affymetrix YG-S98 GeneChips® was performed as described in the Affymetrix user’s manual. Data acquisition was performed using the software packages Microarray Suite v5.0, MicroDB v3.0 and Data Mining Tool v3.0 (Affymetrix, Santa Clara, CA, USA).
Hybridization protocol
One hundred millilitres of culture was sampled directly from the fermentor into a beaker containing 200–300 ml of liquid nitrogen and then processed . Total RNA was extracted with phenol/chloroform. mRNA extraction, cDNA synthesis and labelling, as well as array hybridisation against Affymetrix YG-S98 GeneChips® was performed as described in the Affymetrix user’s manual. Data acquisition was performed using the software packages Microarray Suite v5.0, MicroDB v3.0 and Data Mining Tool v3.0 (Affymetrix, Santa Clara, CA, USA).
Scan protocol
One hundred millilitres of culture was sampled directly from the fermentor into a beaker containing 200–300 ml of liquid nitrogen and then processed . Total RNA was extracted with phenol/chloroform. mRNA extraction, cDNA synthesis and labelling, as well as array hybridisation against Affymetrix YG-S98 GeneChips® was performed as described in the Affymetrix user’s manual. Data acquisition was performed using the software packages Microarray Suite v5.0, MicroDB v3.0 and Data Mining Tool v3.0 (Affymetrix, Santa Clara, CA, USA).
Description
J09
Data processing
MAS5.0 calculated intensity with global array targetting at 150
