Affymetrix GeneChip Yeast Genome 2.0 Array [Yeast_2]

Mapping Condition-Dependent Regulation of Lipid Metabolism in Saccharomyces cerevisiae.

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Investigation of lipid metabolism as a function of three experimental factors: 1) aerobicity (aerobic vs anaerobic, or 'A' vs 'O'), 2) nutrient limitation (carbon vs nitrogen limitation, or 'C' vs 'N'), 3) temperature (30C vs 15C, or 'T' vs 't') using a full factorial design.

GROWTH CONDITION_1

GROWTH CONDITION_2

TEMPERATURE

Transcription profiling by array of condition dependent regulation of lipid metabolism in Saccharomyces cerevisiae

Investigation of lipid metabolism as a function of three experimental factors: 1) aerobicity (aerobic vs anaerobic, or 'A' vs 'O'), 2) nutrient limitation (carbon vs nitrogen limitation, or 'C' vs 'N'), 3) temperature (30C vs 15C, or 'T' vs 't') using a full factorial design.


P-MTAB-27948

Steady state aerobic chemostat cultures were grown at 15C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 25 g glucose, 5 g (NH4)2SO4, 3 g KH2PO4, 0.5 g MgSO4·7H2O, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Aerobic conditions were maintained by sparging with air (1.0 L min-1).(Parameters: time unit = seconds, min temperature = 15, temperature unit = C, media = Minimal media, C-limited)

P-MTAB-27949

Steady state aerobic chemostat cultures were grown at 30C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 25 g glucose, 5 g (NH4)2SO4, 3 g KH2PO4, 0.5 g MgSO4·7H2O, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Aerobic conditions were maintained by sparging with air (1.0 L min-1).(Parameters: time unit = seconds, min temperature = 30, temperature unit = C, media = Minimal media, C-limited)

P-MTAB-27950

Steady state anaerobic chemostat cultures were grown at 30C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 74 g glucose, 1 g (NH4)2SO4, 3 g KH2PO4, 5.3 g K2SO4, 0.5 g MgSO4·7H2O, 420 mg Tween 80, 10 mg ergosterol, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Anaerobic conditions were maintained by sparging the medium reservoir and the fermentor with pure nitrogen gas (0.5 L/min).(Parameters: time unit = seconds, min temperature = 30, temperature unit = C, media = minimal media, N-limited)

P-MTAB-27951

Steady state anaerobic chemostat cultures were grown at 15C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 46 g glucose, 1 g (NH4)2SO4, 3 g KH2PO4, 5.3 g K2SO4, 0.5 g MgSO4·7H2O, 420 mg Tween 80, 10 mg ergosterol, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Anaerobic conditions were maintained by sparging the medium reservoir and the fermentor with pure nitrogen gas (0.5 L/min).(Parameters: time unit = seconds, min temperature = 15, temperature unit = C, media = minimal media, N-limited)

P-MTAB-27952

Steady state aerobic chemostat cultures were grown at 15C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 46 g glucose, 1 g (NH4)2SO4, 3 g KH2PO4, 5.3 g K2SO4, 0.5 g MgSO4·7H2O, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Aerobic conditions were maintained by sparging the cultures with air (1.0 L min-1).(Parameters: time unit = seconds, min temperature = 15, temperature unit = C, media = minimal media, N-limited)

P-MTAB-27953

Steady state anaerobic chemostat cultures were grown at 15C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 25 g glucose, 5 g (NH4)2SO4, 3 g KH2PO4, 0.5 g MgSO4·7H2O, 420 mg Tween 80, 10 mg ergosterol, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Anaerobic conditions were maintained by sparging the medium reservoir and the fermentor with pure nitrogen gas (0.5 L/min).(Parameters: time unit = seconds, min temperature = 15, temperature unit = C, media = Minimal media, C-limited)

P-MTAB-27954

40 – 60 ml of culture broth was sampled directly from the chemostat into a beaker containing 200 ml of liquid nitrogen. The frozen sample was then broken into small fragments and transferred to a 50-ml centrifuge tube. The sample was then thawed at room temperature, ensuring that it remained as close to zero as possible. Cells were pelleted (5000 rpm at 0 C for 4 min), re-suspended in 2 ml of ice-cold AE buffer (50 mM sodium acetate, 10 mM EDTA, pH 5.0) and aliquoted into 5 Eppendorf tubes. This corresponded to 20 mg of dry weight per tube. Total RNA was extracted using FastRNA Pro RED kit (QBiogene, Inc, USA) according to manufacturer's instructions.(Parameters: Extracted product = total_RNA, Amplification = none)

P-MTAB-27955

Washing and staining of arrays were performed using the GeneChip Fluidics Station 450 and scanning with the Affymetrix Gene Array Scanner 3000 7G. Affymetrix Microarray Suite v5.0 was used to generate CEL files of the scanned DNA microarrays.(Parameters: Scanning hardware =  GeneChip 3000 7G Scanner [Affymetrix], Scanning software = Scanning software)

P-MTAB-27956

Steady state aerobic chemostat cultures were grown at 30C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 74 g glucose, 1 g (NH4)2SO4, 3 g KH2PO4, 5.3 g K2SO4, 0.5 g MgSO4·7H2O, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Aerobic conditions were maintained by sparging with air (1.0 L min-1).(Parameters: time unit = seconds, min temperature = 30, temperature unit = C, media = minimal media, N-limited)

P-MTAB-27957

P-MTAB-27958

Steady state anaerobic chemostat cultures were grown at 30C in 2 L jacketed bioreactors (Braun Biotech) using a working volume of 0.5 L and a dilution rate of D = 0.050 (± 0.005) h-1. Cultures were fed with a modified minimal medium containing (amounts per liter): 25 g glucose, 5 g (NH4)2SO4, 3 g KH2PO4, 0.5 g MgSO4·7H2O, 420 mg Tween 80, 10 mg ergosterol, 1 mL Trace Metal Solution, 0.05 mL antifoaming agent and 1 mL vitamin solution. Anaerobic conditions were maintained by sparging the medium reservoir and the fermentor with pure nitrogen gas (0.5 L/min).(Parameters: time unit = seconds, min temperature = 30, temperature unit = C, media = Minimal media, C-limited)

P-MTAB-27959

20 mL of culture was placed into a tube with 35-40 mL of crushed ice in order to decrease the sample temperature to below 2 C in less than 10 seconds.  Cells were then centrifuged (4500 rpm at 0 C for 3 minutes), instantly frozen in liquid nitrogen and stored at -80 C. Total RNA was extracted using FastRNA Pro RED kit (QBiogene, Inc, USA) according to manufacturer's instruction.(Parameters: Extracted product = total_RNA, Amplification = none)

Transcription profiling by array of condition dependent regulation of lipid metabolism in Saccharomyces cerevisiae

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