GSM6008369: S34 at L-10; Homo sapiens; RNA-Seq
GSM6008369
sEVs were isolated from peripheral blood (PB) samples of using the ExoQuick Plasma preparation and sEV precipitation kit (Cat # EXOQ5TM, System Biosciences, CA, USA). Samples were subsequently centrifuged at 10,000 rpm for 5 minutes, incubated with the exosome precipitation solution and refrigerated at 4ºC for 30 minutes. After centrifugation at 1,500 x g for 30 minutes at 4ºC, a beige-colored pellet was dissolved in sterile 1xPBS. Total RNA from sEVs was isolated using miRNeasy kit following manufacturer's protocol (Cat#217084, Qiagen). RNA sample quality was assessed by NanoDrop and Agilent 2100 BioAnalyzer. Samples with RNA integrity number (RIN) above 7, OD260/280: 2, and OD260/230 ≥ 2 were used. In addition, RNA degradation and contamination were monitored on 1% agarose gels before RNA-Seq. RNA quality was assessed using an Agilent TapeStation (Agilent, Palo Alto, CA, USA), and RNA concentration was quantified by Qubit 4.0 spectrophotometer. The library for small RNA sequencing was prepared using the Smarter smRNA-seq kit for Illumina (Takara Bio Inc., USA). The quantity and quality of amplified libraries were evaluated using Qubit (Invitrogen, Carlsbad, CA, USA) and Agilent TapeStation high sensitivity D1000 Screen Tape. Small RNA-seq libraries were sequenced using single-end 75 base pairs (SE75) sequencing chemistry on NextSeq 500 instruments following the manufacturer's protocols (Illumina).