characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2020-09-02T04:06:54Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2019-10-15T16:11:09Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA6086355
characteristics.characteristic: External Id
characteristics.text: Microbial Genomics and Biotechnology, Center for Biotechnology, Bielefeld University
characteristics.characteristic: INSDC center name
characteristics.text: 2020-09-02T04:06:54Z
characteristics.characteristic: INSDC first public
characteristics.text: 2019-10-15T16:11:09Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS3886763
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-8432:wt growth phase
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: Protocols: For transcriptome analysis, 2x 1 mL samples from Actinoplanes culture were taken during exponential growth phase, separated from the supernatant by centrifugation (10 s) and snap-frozen in liquid nitrogen. Pellets were stored at -80 °C until further processing. Cultivation was performed in 250 mL Corning® Erlenmeyer baffled cell culture flasks at 28 °C and 140 rpm for seven days in maltose minimal medium (72.06 g?L-1 maltose·1H2O, 5 g?L-1 (NH4)2SO4, 0.184 g?L-1 FeCl2·4H2O, 5.7 g?L-1 Na3C6H5O7·2H2O, 1 g?L-1 MgCl2·6H2O, 2 g?L-1 CaCl2·2H2O, trace elements (final concentration: 1 µM CuCl2, 50 µM ZnCl2, 7.5 µM MnCl2 dissolved in 1 M HCl) and phosphate buffer consisting of 5 g?L-1 each K2HPO4 and KH2PO4 in aqua distilled). The medium was prepared and filter sterilized following the protocol of Wendler et al. (2013). For inoculation of 50 mL medium, 1 mL spore solution of an OD = 3-5 was used. Cell dry weights were determined like described by Wolf et al. (2017). The supernatant was stored for later analysis at -20 °C. For RNA isolation, frozen cell pellets were resuspended in 500 µL LB-buffer (NucleoSpin® RNA Plus, Macherey-Nagel, Düren, Germany) and transferred to 2 mL lysing matrix tubes (0.1 mm spherical silica beads, MP Biomedicals, Santa Ana, California, USA). Cell disruption was carried out in a homogenizer (FastPrep FP120, Thermo Fisher Scientific, Waltham, MA, USA) for three times 20 s at speed setting 6.5 and 5 min on ice in between. Subsequently, the cell suspension was centrifuged for 5 min at 13,000 g and 4 °C. The supernatant was used for RNA extraction using the NucleoSpin® RNA Plus Kit in combination with rDNase Set (Macherey-Nagel, Düren, Germany) for an on-column DNA digestion. After clean-up and elution according to manufacturer's protocol, the DNA-digestion was repeated in-solution and the sample cleaned up again by use of the same kit. Residual DNA was tested negatively with two primer pairs binding to genomic DNA of Actinoplanes sp. SE50/110 and amplifying small fragments at round about 200-300 nt. The quantity of RNA was analyzed with the NanoDrop 1000 spectrometer (Peqlab, Erlangen, Germany). RNA of triplicates of the wildtype and the deletion mutant was isolated from growing cultures and pooled equimolar separately for the wildtype and the deletion mutant (total amount of 2.5 µg RNA each in 26 µL RNase-free water). rRNA was depleted using the Ribo-Zero rRNA Removal Kit for bacteria (Illumina, San Diego, USA). Successful depletion of rRNA was verified by an Agilent RNA 6000 Pico chip in the Bioanalyzer (Agilent, Böblingen, Germany). cDNA libraries were prepared following protocols from Pfeifer-Sancar et al. (2013) and Irla et al. (2015) by use of the TruSeq stranded mRNA kit (Illumina, San Diego, CA, USA). The libraries were quantified by a DNA High Sensitivity Assay chip in the Bioanalyzer (Agilent, Böblingen, Germany).
characteristics.characteristic: description
characteristics.text: wild type
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005168
characteristics.characteristic: genotype
characteristics.text: growth phase
characteristics.characteristic: growth condition
characteristics.text: Actinoplanes sp. SE50/110
characteristics.characteristic: organism
characteristics.text: Actinoplanes sp. SE50/110
characteristics.characteristic: organism
characteristics.text: Actinoplanes sp. SE50/110
characteristics.characteristic: scientific_name
characteristics.text: wt growth phase
characteristics.characteristic: title