title: GSM8978214: Patient B5, severe sepsis originating from urogenital infection, miRNA; Homo sapiens; miRNA-Seq
library_construction_protocol: RNAs from blood samples were isolated with TRIzol protocol and Tempus™ Spin RNA Isolation Kit according to the manufacturers' instructions (TRIzol™ Reagent User Guide1 – ThermoFisher Scientific, Inc., Waltham, MA, USA; Tempus Blood RNA Tube and Tempus Spin RNA Isolation Kit User Guide2 ThermoFisher Scientific, Inc., Waltham, MA, USA). RNA concentration, quality and RNA integrity number (RIN) of each sample were measured using Bioanalyzer 2100 (Agilent Technologies, Inc., Santa Clara, CA, USA) instruments using Eukaryotic Total RNA Nano Kit (RNA 6000) (Agilent Technologies, Inc., Santa Clara, CA, USA) according to manufacturer's protocol. RNA-Seq libraries were prepared from total RNA using Ultra II RNA Sample Prep kit (New England BioLabs, Inc., Ipswich, MA, USA) according to the manufacturer's protocol. Briefly, poly-A RNAs were captured by oligo-dT conjugated magnetic beads then the mRNAs were eluted and fragmented at 94-Celsius degree. First strand cDNA was generated by random priming reverse transcription and after second strand synthesis step double stranded cDNA was generated. After repairing ends, A-tailing and adapter ligation steps adapter ligated fragments were amplified in enrichment PCR and finally sequencing libraries were generated.