title: GSM4200699: S759N-1; Escherichia coli K-12; OTHER
library_construction_protocol: HydEn-seq method has been previously described. Briefly, DNA extraction was performed using MasterPure™ Complete DNA and RNA Purification Kit (Lucigen, Epicentre, MC85200). After measuring the DNA concentration using Qubit® dsDNA BR Assay Kit (Life Technologies, molecular probes, Q32853), 1 µg DNA was hydrolyzed using 0.3 M KOH by incubation at 55°C for 2 hours. Ethanol-precipitated DNA was denatured by 3 minutes incubation at 85°C. The single strand DNA containing 5´-OH ends was phosphorylated using 3´-phosphatase minus T4 polynucleotide kinase (BioLabs, M0236L), incubated at 65°C for 20 min, washed with beads (GC biotech, CPCR-0050), denatured, ligated to ARC140 adaptor using T4 RNA Ligase 1 (BioLabs, M0204S), denatured, polymerized using T7 DNA Polymerase (BioLabs, M0274L) and ARC76/77 oligos, and bead-washed. KAPA HiFi hotstart readymix (2X) kit (KAPA biosystems, 07958935001) was used to amplify the libraries. Paired-end sequencing was performed using Illumina NextSeq 500 system.
attributes.tag: GEO Accession
attributes.value: GSM4200699