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Sample GSM1354499

Query DataSets for GSM1354499

Status

Public on Jul 16, 2014

Title

MV01_Kar18L

Sample type

SRA

Source name

Aerobic batch fermentor

Organism

Kluyveromyces lactis

Characteristics

strain: GG1632
genotype/variation: Klku80Δ KlCAR; reference strain
culture condition: aerobic bioreactor batch cultures on glucose chemically defined medium with arginine as sole nitrogen source

Treatment protocol

Broth was directly quenched in liquid nitrogen.

Growth protocol

Growth experiments were conducted in synthetic medium containing salts, trace elements and vitamins, prepared and sterilized as described previously (Verduyn et al., 1992). Glucose was added to a final concentration of 20.0 g L-1 and 3.3g/l arginine was used. Moreover, 3.3 g L-1 potassium sulfate was added to compensate for the removal of ammonium sulfate. Controlled aerobic batch cultures were grown in 2 L laboratory bioreactors (Applikon, Schiedam, The Netherlands) with a working volume of 1.2 L. Culture pH was maintained at pH 6.0 by automatic addition of 2 M KOH (ADI 1030 Biocontroller, Applikon) and temperature was kept constant at 30C (Lauda E300 thermostat, Lauda DR. R. Wosber Gmbh & Co. KG Lauda-Königshofen, Germany). Aeration was achieved by sparging air through the cultures at a flow rate of 0.5 L min-1 continuously stirring at 700 rpm. The exhaust gas of the bioreactor was cooled in a condenser using a cryostat set at 4C (Lauda E100 cryostat). CO2 and O2 concentrations in the off gas were measured with an NGA 2000 analyzer (Rosemount Analytical, Solon, OH).

Extracted molecule

total RNA

Extraction protocol

Biomass samples for RNA extractions were sampled from mid-exponential phase of K. lactis strains GG and IMS0367 grown in batch cultivations in bioreactors with chemically defined medium with glucose and arginine as sole N-source. The sampling, the extract
cDNA libraries for the Illumina platform were generated and sequenced at BaseClear BV (Leiden, The Netherlands). cDNA was used as input for library preparation using the Illumina TruSeq DNA library preparation kit (Illumina). Briefly, the DNA was fragmented by nebulization and subjected to end-repair, A-tailing, ligation of adaptors including sample-specific barcodes and size-selection to obtain a library with median insert-size around 200-400 bp. After PCR enrichment, the resultant library was checked on a Bioanalyzer (Agilent) and quantified. The libraries were multiplexed, clustered, and sequenced on an Illumina HiSeq 2500 with paired-end protocol
library with a 350 insert size were prepared and sequenced with an Illumina Genome Analyzer II.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina Genome Analyzer II

Description

Sample 1

Data processing

At Baseclear FASTQ sequence reads were generated using Illumina Casava pipeline version 1.8.2
Quality assesment was based on data passing the Ilumina Chastity filtering. Subsequently, reads containing adapters and/or PhiX control signal were removed using an in-house (At Baseclear) filtering protocol.
RNA-seq reads were mapped with BWA (version 0.5.9-r16) to Kluyveromyces lactis NRRL Y-1140 whole genome
samtools (version 0.1.7-r510) was used to convert the BWA sam output to sorted bam format
cuffdiff, from the cufflinks packages (version 1.0.3), was used to calculate the differential expression between WT (MV01_Kar18L and _18R) and Mutant (MV01_Kar9L and _Kar9R).
Genome_build: Kluyveromyces lactis strain CLIB210/NRRL Y-1140 GCF_000002515.2
Supplementary_files_format_and_content: differential expression between WT and Mutant

Submission date

Mar 20, 2014

Last update date

May 15, 2019

Contact name

Jean-Marc Daran

E-mail(s)

j.g.daran@tudelft.nl

Phone

+31 15 278 2412

Organization name

Delft University of Technology