miR-seq of 100K-EVs isolated from BV-2 cells traited with WT-GAGs - rep 2
Total RNAs from EV preparations and 0.5  106 cells were extracted using the miRNeasy Tissue/Cells Advanced Micro Kit (QIAGEN), according to the manufacturers instructions. The purified RNAs were eluted into 20 l nuclease-free water, and stored at  80 C until analysis. RNA was quantified using the Qubit RNA HS Assay Kit on the Qubit Fluorometer (Invitrogen). The quality was evaluated on an Agilent 4150 TapeStation with the High Sensibility RNA ScreenTape kit (Agilent). Each samples libraries were built with the QIAseq miRNA UDI Library Kit (QIAGEN), according to the manufacturers instructions and quantified using the Qubit 1X dsDNA HS kit on the Qubit Fluorometer (Invitrogen). All libraries were sequenced using a single-read strategy (10 million reads per sample) on an NextSeq550 platform, with 150 cycles of the MidOutput flowcell (Illumina), a read length of 72 bp, and dual indices of 10 bp. Raw data were converted to FASTQ files using bcl2fastq (v2.20).