title: GSM3983616: C3-2; Vibrio cholerae O1 biovar El Tor str. N16961; RNA-Seq
library_construction_protocol: RNA was isolated by using TRIzol reagent (Ambion), breifly the pellet resuspended and left at ambient temperature for 5 min. Next, 260 µL of chloroform was added, followed by shaking and centrifugation (11,500 × g, 5 min). The upper (aqueous) phase was transferred to a fresh tube, mixed with an equal volume of chloroform, and centrifuged. The upper phase was then transferred to 600 µL of isopropanol, vortexed, and incubated at room temperature for 10 min, followed by centrifugation (12,000 × g, 15 min, 4 °C). The pellet (total RNA) was washed twice with 75% (vol/vol) ethanol and then resuspended in 100 µL of nuclease-free water (Ambion). DNA was removed using the Turbo DNase kit (Ambion). RNA samples were quantified using Qubit 2.0 Fluorometer (Life Technologies), and the RNA integrity was checked with RNA6000 Nano Assay using the Agilent 2100 Bioanalyzer (Agilent Technologies). cDNA library preparation and sequencing reactions were conducted by Genewiz. Illumina TruSeq Stranded Total RNA Library Prep Kit, clustering, and sequencing reagents were used throughout the process following the manufacturer's recommendations. The cDNA library was quantified using Qubit 2.0 Fluorometer (Life Technologies) and by qPCR. The samples were clustered on a flow cell using the cBOT. After clustering, the samples were loaded for sequencing of 1 × 50 single read on the Illumina HiSEq. 2500 instrument.
attributes.tag: GEO Accession
attributes.value: GSM3983616