Novosphingobium resinovorum
Sample type
SRA
strain
SA1
source name
SA1, Glc, Stat, Rep 1
medium
10 mM glucose (Glc) , 10 mM NH4Cl
growth phase
stationry (Stat)
Growth protocol
N. resinovorum SA1 strain was grown in 250 mL Erlenmeyer flasks in 100 mL minimal salt media (MM) according Perei et al. (1), containing 10 mM substrates (either sulfanilic acid (SA), or glucose (Glc)), 25 µg/ml (Sm) and 25 mM MOPS (pH=7.0). Addition of MOPS could stabilize the pH during the cell growth, the pH change/drop was less than 0.2 unit in any cultivation. When glucose was used as carbon source, the medium was supplemented with 10 mM NH4Cl, in the case of SA, the substrate was the sole nitrogen source. The cultures were shaken at 28 °C, 180 rpm and the cell growth was monitored by measuring the OD600nm values. Samples for RNA extraction were taken from exponential and late stationary phase. 1. Perei K, Rákhely G, Kiss I, Polyák B, Kovács KL. Biodegradation of sulfanilic acid by Pseudomonas paucimobilis. Appl Microbiol Biotechnol. 2001;55(1):101-107. http://www.ncbi.nlm.nih.gov/pubmed/11234949
Extracted molecule
total RNA
Extraction protocol
For RNA-Seq analysis, 6 ml samples were taken from exponential and late stationary phase cultures grown on Glc or SA in MM. In each case, two independent biological replicas were used. The samples were harvested (17,226 rcf, 5 min, 4 °C) and immediately frozen in liquid nitrogen and stored at -80°C until RNA isolation. The frozen samples were resuspended in 350 µl Zymo DNA/RNA Shield. The resuspended cells were mixed with 700 µl RLT buffer (Qiagen) (with ?-merkaptoetanol) and were transferred into a new tube containing 0.8 g, 0.5 mm Glass Beads (Scientific Industries, SI-BG05). The cells were vortexed for 5 minutes on maximum speed with a Vortex-Genie 2 (Scientific Industries, SI-0236) supplemented with a TurboMix Attachment (Scientific Industries, SI-0564). Following lysis, the mixtures were centrifuged at 9,600 rcf for 10 seconds to remove the glass beads and cell debris. The supernatants were used for purification of total RNAs with the Qiagen RNeasy Mini Kit protocol (Qiagen) with on-column DNase digestion. Then the Purified RNAs were treated again with RNase free DNase I (Sigma) to remove any residual genomic DNA contamination. Finally, the RNAs obtained were repurified with the RNeasy Mini Kit Clean Up protocol (Qiagen). The integrity of the RNA was checked by Agilent 2100 Bioanalyzer. RNA libraries were prepared for sequencing using standard Illumina epicenter ScriptSeq (Bacteria) protocol.
collection_date
missing
geo_loc_name
missing
sample_type
total RNA
host
missing
BioSampleModel
Microbe, viral or environmental