alias: GSM4872575
title: GSM4872575: Islet-derived exosomes-Control-Sample 7; Homo sapiens; miRNA-Seq
library_construction_protocol: Untreated and cytokine treated human islets and islet-cell supernatants were harvested for downstream applications. For exosome isolation, supernatants were collected and centrifuged for 30 minutes at 3000g to remove cellular debris. Exosomes were isolated using ExoQuick-TC (SBI Bioscience, USA) as per the manufacturer's instructions. Total RNA and miRNA were first evaluated for quantity, quality, and percent miRNA content in total RNA, using the Agilent Bioanalyzer. The starting amount of RNA was 10-20 ng for each miRNA library. For miRNA library preparation, 0.5% of miRNA in total RNA was the cut-off to decide whether the sample should be enriched for miRNA. When needed, enrichment was performed following the small RNA library preparation procedure and the Ion Total RNA-Seq Kit v2 User Guide, Pub. No. 4476286 Rev. E (Life Technologies, USA). Each resulting barcoded library was quantified and assessed for quality using the Agilent Bioanalyzer. Multiple libraries were pooled in equal molarity. Eight microliters of 100 pM pooled libraries were applied to Ion Sphere Particles (ISP) template preparation and amplification using Ion OneTouch 2, followed by ISP loading onto PI chips and sequencing using Ion Proton semiconductor. Each PI chip is capable of generating approximately 50 million usable reads of 21-22bp miRNA fragments. Sequence mapping was performed using Torrent Suite Software v4.6 (TSS 4.6).
identifiers.namespace: GEO
identifiers.id: GSM4872575
attributes.tag: GEO Accession
attributes.value: GSM4872575