GSM4988932: Normal3; Homo sapiens; miRNA-Seq
The ExoQuick kit (System Biosciences, Mountain View, CA) was applied to isolate exosomes as according to the manufacturer's protocol. Frozen plasma was thawed and centrifuged at 21,000 g for fifteen min at 4°C to purify them. The supernatant was later moved to a clean tube, and quarter of EXOQ5A-1 solution was added and mixed lightly. After 2 hours incubating at 4°C, the mix was centrifuged at 1,500 g for 30 min, and the supernatant was removed using aspiration. The residual supernatant was removed after centrifugation for 5 minutes; exosome containing pellets were resuspended in water free of nuclease or relevant buffer for RNA and protein extraction. The entire 3 mg total RNA of each sample was divided into aliquots and a small RNA (sRNA) library prepared. The NEBNext Multiplex sRNA Library Prep Set for Illumina (New England Biolabs, Ipswich, MA, USA) was applied to generate the sequencing library. The index code was added to assign a sequence to every specimen. TruSeq SR Cluster Kit v3-cBot-HS (Illumina, San Diego, CA, USA) was applied to cluster index-coded specimens. The libraries were sequenced on an Illumina Hiseq 2500 machine to generate the 50 bp single-end reads.
GEO
GSM4988932