title: Illumina HiSeq 2000 sequencing; The core genome m5C methyltransferase JHP1050 (M.Hpy99III) plays an important role in orchestrating gene expression in Helicobacter pylori
design: The core genome m5C methyltransferase JHP1050 (M.Hpy99III) plays an important role in orchestrating gene expression in Helicobacter pylori
library_construction_protocol: H. pylori strains were grown in liquid cultures as described in: Bubendorfer, S., Krebes, J., Yang, I., Hage, E., Schulz, T.F., Bahlawane, C., Didelot, X. and Suerbaum, S. (2016) Genome-wide analysis of chromosomal import patterns after natural transformation of Helicobacter pylori. Nat. Commun., 7, 11995.  For RNA extraction, 5 ml of bacterial cells grown in liquid medium were pelleted (4°C, 6000 x g, 3 110 min), snap-frozen in liquid nitrogen and stored at -80 °C. Afterwards, bacterial pellets were disrupted with a FastPrep® FP120 Cell Disrupter (Thermo Savant) using Lysing Matrix B 2 ml tubes containing 0.1 mm silica beads (MP Biomedicals, Eschwege, Germany). Isolation of RNA was performed using the RNeasy kit (Qiagen, Hilden, Germany) and on-column DNase digestion with DNase I. A second DNase treatment was carried out using the TURBO DNA-free™ Kit (Ambion, Kaufungen, Germany).  Total rRNA depletion was performed prior to cDNA synthesis using a RiboZero Kit (Illumina, Germany). cDNA library was performed with random primers.
attributes.tag: Experimental Factor: strain
attributes.value: J99
attributes.tag: Experimental Factor: phenotype
attributes.value: GCGC-MTase JHP1050 mutant