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| Status |
Public on Jul 21, 2018 |
| Title |
S. eubayanus CBS12357-1 |
| Sample type |
SRA |
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| Source name |
cultures
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| Organism |
Saccharomyces eubayanus |
| Characteristics |
carbon source: Glucose
strain: CBS 12357
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| Treatment protocol |
S. eubayanus CBS 12357T , S. kudriavzevii and S. cerevisiae Culture samples corresponding to ca. 240 mg of biomass wet weight were directly quenched in liquid nitrogen.
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| Growth protocol |
S. eubayanus, S. kudriavzevii and S. cerevisiae batch cultures were grown in bioreactors performed in independent duplicate. SMG was supplemented with 0.,2 g L -1 antifoam Emulsion C (Sigma-Aldrich, St. Louis, MO). The reactors were inoculated at OD660 0.,3 with cells resuspended in demineralized water, which were obtained from exponential growing shake flask cultures growing at the same temperature and medium as was used in the bioreactors. Cultures were performed in 2 L bioreactors (Applikon, Schiedam, The Netherlands) containing a 1.,4 L working volume. The cultures were constantly stirred at 800 rpm, sparged with 700 mL min-1 dried compressed air (Linde Gas Benelux, The Netherlands) and maintained at 30 ˚C for S. cerevisiae and 25 ˚C for S. kudriavzevii and S. eubayanus. The culture pH was kept at 5.0 during growth on glucose by automatic addition of 2M KOH. (SM) containing 3.0 g L-1 KH2PO4, 5.0 g L-1 (NH4)2SO4, 0.5 g L-1 MgSO4, 7 H2O, 1 mL L-1 trace element solution, and 1 mL L-1 vitamin solution. The pH was set to 6 with 2 M KOH prior to autoclaving at 120 °C for 20 min. Vitamin solutions (Verduyn et al. 1992) were sterilized by filtration and added to the sterile medium. Concentrated sugar solutions were autoclaved at 110 °C for 20 min and added to the sterile flasks to give a final concentration of 20 g L-1 carbon source (glucose (SMG), maltose (SMM)
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
The resulting ice pellet was gently thawed on ice and spun down at 4700 x g for 5 min at 0 o C. Pellets were then resuspended in 1.2 mL of ice-cold AE buffer (50 mM sodium acetate and 10 mM EDTA, pH 5.0), followed by addition of 1.2 mL of acid phenol/chloroform/isoamyl alcohol mix and 0.12 mL 10 % sodium dodecyl sulfate. The resulting mix was vortexed for 30 s and incubated for 5 min at 65 °C. After homogenizing for 30 sec by vortexing, 800 µL aliquots were distributed in RNase-free screw-cap tubes (Tai et al. 2005). After centrifugation (15 min at 10,000 x g), the aqueous phase was transferred to a new tube containing 0.4 mL of acid phenol/chloroform. The mix was vortexed for 30 seconds, centrifuged (15 min at 10,000 x g) and the aqueous phase was again transferred to a new tube. RNA was then ethanol precipitated and re-suspended in RNAse-free water. Prior to cDNA synthesis, purity, concentration and integrity of the RNA in the samples was assessed with the Nanodrop (Thermo Fisher Scientific), Qubit (Thermo Fisher Scientific) and Tapestation 220 with RNA Screen Tape (Agilent Technologies), respectively, according the manufacturers’ recommendations.
cDNA libraries were prepared using the TruSeq RNA V2 kit (Illumina)and sequenced on HISeq 2500 (Illumina) at Novogene (HK) Company Ltd (Hong Kong, China).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
S. eubayanus CBS 12357T was grown in in 2 L bioreactors (Applikon, Schiedam, The Netherlands) containing a 1.,4 L working volume. The cultures were constantly stirred at 800 rpm, sparged with 700 mL min-1 dried compressed air (Linde Gas Benelux, The Netherlands) and maintained at 30 ˚C for S. cerevisiae and 25 ˚C for S. kudriavzevii and S. eubayanus. The culture pH was kept at 5.0 during growth on glucose by automatic addition of 2M KOH. . Culture samples corresponding to ca. 240 mg of biomass wet weight were directly quenched in liquid nitrogen.
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| Data processing |
RNA-Seq reads were aligned to S. eubayanus CBS12357T, S. kudriavzevii CR85 and S. cerevisiae CEN.PK113-7D reference using STAR version 2.5.3a in 2Pass mode
resulted sequence alignment map file (SAM) were filtered on ambigious alignments and sorted with samtools to sorted binary alignement file (BAM)
The sorted BAM file was processed with htseq-count from the HTSeq framework version 0.9.1 with -m union and -stranded reverse to calculate the transcript counts
The raw transcript counts were normalized on fragments per kilobase million (FPKM) by applying rpkm module from edgeR package
Genome_build: S. eubayanus CBS12357 from Baker et al. for sample 1 and 2; S. kudriavzevii CR85 from Boonekamp et al. for samples 3 and 4; S. cerevisiae CEN.PK113-7D from Salazar et al. for samples 5 and 6
Supplementary_files_format_and_content: Comma delimited file, normalized transcript counts
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| Submission date |
Jul 19, 2018 |
| Last update date |
Jul 22, 2018 |
| Contact name |
Jean-Marc Daran |
| E-mail(s) |
j.g.daran@tudelft.nl
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| Phone |
+31 15 278 2412
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| Organization name |
Delft University of Technology
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| Department |
Department of Biotechnology
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| Lab |
Kluyver centre for genomics of industrial organisms
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| Street address |
Julianalaan 67
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| City |
Delft |
| ZIP/Postal code |
2628BC |
| Country |
Netherlands |
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| Platform ID |
GPL25341 |
| Series (1) |
| GSE117404 |
Exploration of the genetic makeup and expression of the glycolytic and fermentative pathways within the Saccharomyces genus |
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| Relations |
| BioSample |
SAMN09695467 |
| SRA |
SRX4411565 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3293891_fpkm_normalized_CBS12357_R5.csv.gz |
168.8 Kb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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