Transcriptomics

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RNA-Seq of Corynebacterium glutamicum ATCC13032 differing in growth rates reveals the growth rate modulon


ABSTRACT: The growth rate (µ) of microbes is a fundamental property influencing its production capacity. To identify the transcriptomic changes of Corynebacterium glutamicum ATCC13032 with increasing growth rate, three transitions, induced by different pre-culture conditions, were sampled in triplicate at growth rates ranging from 0.02 to 0.4 h-1. The pre-culture conditions differed in limiting substrate (phosphate, nitrogen, carbon) and the length of the stationary phase. Samples of 2 mL were withdrawn from the bioreactor in biological triplicates and immediately centrifuged at 20000 g for 30 seconds at 4 °C. The supernatant was discarded and the remaining cell pellet was immediately flash frozen in liquid nitrogen. Total RNA was isolated from three biological replicates using RNeasy Mini Kit along with a DNase Kit (both from Qiagen). Initially RNA quality was checked by Trinean Xpose (Gentbrugge,Belgium) and Agilent RNA Nano 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). Samples contaminated with DNA were treated with DNase (Qiagen), cleaned as described above and rechecked by Xpose and Agilent Bioanalyzer. Finally RNA was free of DNA with an RNA Integrity Number (RIN) > 9 and rRNA Ratio [23s / 16s] > 1.5. Ribo-Zero rRNA Removal Kit (Bacteria) from Illumina (San Diego, CA, USA) was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was checked by Agilent RNA Pico 6000 kit on Agilent 2100 Bioanalyzer (Agilent Technologies, Böblingen, Germany). RNA was free of detectable rRNA. TruSeq Stranded mRNA Library Prep Kit from Illumina (San Diego, CA, USA) was used to prepare cDNA libraries. The resulting cDNAs were sequenced paired end on an Illumina MiSeq system using 75 bp read length and on Illumina HiSeq 1500 system using 70 bp read length and 50 bp read length for one single sample. Through the comparison of these three datasets, each containing three biological replicates, the pre-condition independent gene expression changes could be deduced and the growth rate modulon was identified.

INSTRUMENT(S): Illumina HiSeq 1500, Illumina MiSeq

ORGANISM(S): Corynebacterium glutamicum ATCC 13032  

SUBMITTER: Thorsten Haas   Ralf Takors   Tobias Busche  

PROVIDER: E-MTAB-7436 | ArrayExpress | 2019-04-01

SECONDARY ACCESSION(S): ERP112375

REPOSITORIES: ArrayExpress, ENA

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