GSM2574064: TSS_TAP-; Streptomyces tsukubensis; OTHER
Total RNA was isolated from obtained cell lysate by phenol-chloroform extraction and ethanol precipitation. Isolated total RNA was incubated at 37oC for 1 h with DNase I (New England Biolabs) to remove genomic DNA. Total RNA was further purified by phenol-chloroform extraction and ethanol precipitation. Quality of the isolated total RNA was checked by visualization using 2% agarose gel electrophoresis. Phosphorylated RNA fragments were purified using RNeasy MinElute Cleanup Kit (Qiagen). Final library construction was performed using NEBNext Multiplex Small RNA Sample Prep Set for Illumina (New England Biolabs) according to manufacturer’s instructions. The amplified library was separated by gel electrophoresis for 120 min at 100 V on 2% SYBR gold (Bio-rad) agarose gel. Final library around 150 bp was size-selected and purified using MinElute Gel extraction kit (Qiagen).