GSM8546105: 20231215-EV-1_BC15; Homo sapiens; RNA-Seq
Peripheral whole blood samples were collected from individuals before therapy or surgery using EDTA-coated vacutainer tubes. Plasma was separated within 2 hr after collection by centrifuge at 1,900 g for 10 min at 4 ℃. All plasma samples were aliquoted and stored at -80°C. Plasma EV were purified by a membrane-affinity approach using exoRNeasy Midi Kit(Qiagen, 77144) following the manufacturer's instructions. Plasma-nonEV (EV-depleted Plasma) component were collected from flow through part of MinElute spin column from kit. Both plasma EV RNA and Plasma nonEV RNA were extracted using QIAzol Lysis Reagent (QIAGEN, Beijing, China) DNA contamination was removed by Recombinant DNase I (RNase-free) (TAKARA, Beijing, China). End repair and polyadenylation were performed using E. coli Poly(A) Polymerase (NEB, Beijing, China), and T4 Polynucleotide Kinase (NEB, Beijing, China). Reverse transcription and libraries were prepared according to DETECTOR-seq protocol with minor modifications by replacing original TSO with uracil-containing TSO and digestion with Uracil-DNA Glycosylase (NEB, Beijing, China) after reverse transcription. Library quantification was performed by Qubit dsDNA HS Kit and fragment size was checked by Agilent 2100 Bioanalyzer. Libraries were sequenced on MGI Tech DNBSEQ-T7 with PE150.