Sample 12
Saccharomyces cerevisiae
Protocols: At an OD600 of 2, samples were removed from the shakeflasks, cooled quickly on ice, and centrifuged at 4 ˚C for 10 mins at 5,000 x g. The supernatant was removed, the pellets were snap-frozen with liquid nitrogen and stored at -80 ˚C. Production media: From Zelle et al (2008 - Malic Acid Production by Saccharomyces cerevisiae: Engineering of Pyruvate Carboxylation, Oxaloacetate Reduction, and Malate Export) 100 g liter−1 glucose, 3 g liter−1 KH2PO4, 6.6 g liter−1 K2SO4, 0.5 g liter−1 MgSO4·7 H2O, and 1 ml trace element solution. After the pH was set to 4.8 with KOH and the solution was heat sterilized for 20 min at 110°C, a filter-sterilized vitamin solution (1 ml liter−1) and urea (to a final concentration of 1 g liter−1) were added. Trace metals per litre - 4.5 g CaCl2·2H2O, 4.5 g ZnSO4·7H2O, 3 g FeSO4·7H2O, 1 g H3BO3, 1 g MnCl2·4H2O, 0.4 g Na2MoO4·2H2O, 0.3 g CoCl2·6H2O, 0.1 g CuSO4·5H2O, 0.1 g KI, 15 g EDTA Vitamins per litre - biotin, 0.05 mg; calcium pantothenate, 1 mg; nicotinic acid, 1 mg; inositol, 25 mg; thiamine HCl, 1 mg; pyridoxineHCl,1mg; andpara-aminobenzoicacid, 0.2 mg For shake flask fermentation experiments, single colonies of each strain were taken from plates and inoculated into 5 mL of liquid SC medium with appropriate amino acid selection in 12 mL culture tubes and incubated at 30 °C, 250 rpm for 2 days. Shake flasks were prepared as follows; 5 g CaCO3 was added to each 500 mL baffled shake flask before the flasks were autoclaved. Once cool, 100 mL of the sterile batch production medium was added to each shake flask. The prepared flasks were inoculated from the pre-cultures to a starting OD600 of 0.5. Fermentations were carried out at 30 ˚C, and shaking set to 250 rpm. The RNeasy mini kit (Qiagen) was used, followed by treatment with DNase (RNase-Free DNase Set, Qiagen) for the digestion of residual genomic DNA. The sequencing libraries were prepared in triplicates using a TruSeq Sample Preparation kit (Illumina Inc., San Diego CA) and were pooled together prior to sequencing. An average cDNA library size was determined using the Agilent DNA 1000 kit on an Agilent 2100 Bioanalyzer.
ENA-FIRST-PUBLIC
2019-01-31T17:02:09Z
ENA-LAST-UPDATE
2019-01-21T14:03:08Z
External Id
SAMEA5230245
INSDC center name
Novo Nordisk Centre for Biosustainability Danish Technical University
INSDC first public
2019-01-31T17:02:09Z
INSDC last update
2019-01-21T14:03:08Z
INSDC status
public
Submitter Id
E-MTAB-7600:Sample 12
broker name
ArrayExpress
cell line
TAM strain - Delft University
cell type
CEN.PK
common name
baker's yeast
genotype
MATa pdc1- pdc5- pdc6- PYC1PYC2::LEU2 BcBAPATEcYdfG::HIS5 TcPanD::URA3
sample name
E-MTAB-7600:Sample 12
scientific_name
Saccharomyces cerevisiae
strain
SThpdh_2779