title: 42-SET-1
organism: Haemophilus influenzae
description: Protocols: The nontypeable H. influenzae (NTHi) strain GE47 examined in this study was taken from a collection of 46 NTHi strains previously studied (Cherkaoui et al., 2016 PMID: 27756711). In more detail, GE47 was isolated from the sputum of one 75 year-old patient. The inoculum suspension (50 mL) was prepared by picking several colonies from an overnight culture at 37°C on chocolate agar plates with a cotton swab and suspending the colonies in BHI broth supplemented with NAD 2 μg/mL and hemin 10 μg/mL (sBHI) to a density of 0.5 McFarland standards. The inoculum suspension was transferred in 2 sets of ten vials. Two mL of the inoculum suspension by vial. One set was then incubated at 37°C and the other at 42°C. The optical densities (D600) of 1 mL aliquots were plotted against time; and the amount of viable cells was determined at selected time points by plating culture dilutions onto chocolate agar plates, followed by overnight incubation at 37°C. The longest incubation time at 42°C that did not significantly affect cell growth was 3h. Therefore this incubation time was used in all the experiments. All culture incubations were carried out in the presence of 5% CO2. Total RNA was purified from bacterial pellets re-suspended in 1mL of TRIzol (Invitrogen) with 0.1-mm glass/zirconia beads and disrupted in a Bead Beater apparatus. The RNA in the aqueous phase was precipitated with ethanol, rinsed several times and solubilised in 50μL of diethylpyrocarbonate-treated water. RNA libraries were generated using NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB)
attributes.tag: ENA first public
attributes.value: 2019-01-15
attributes.tag: ENA last update
attributes.value: 2017-11-23
attributes.tag: External Id
attributes.value: SAMEA104399875
attributes.tag: INSDC center alias
attributes.value: unige
attributes.tag: INSDC center name
attributes.value: University of Geneva
attributes.tag: INSDC first public
attributes.value: 2019-01-15T17:02:12Z
attributes.tag: INSDC last update
attributes.value: 2017-11-23T11:40:30Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-6237:42-SET-1
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: growth condition
attributes.value: BHI broth supplemented with NAD 2 g/mL and hemin 10 g/mL
attributes.tag: sample name
attributes.value: E-MTAB-6237:42-SET-1
attributes.tag: strain
attributes.value: GE47