GSM7156630: plasma exosomes, osteosarcoma patient [AA230-002]; Homo sapiens; RNA-Seq
a procedure of ultracentrifuged and size exclution chromatography methods  were used to isolate exosomes. Total RNA was extracted and purified from plasma exosome using miRNeasy Serum/Plasma Advanced Kit (Qiagen, cat. No. 217204) according to the kit instruction. RNA concentration and purity were evaluated using the RNA Nano 6000 Assay Kit of the Agilent Bioanalyzer 2100 System (Agilent Technologies, CA, USA). A total amount of 250pg—10ng RNA per sample was used as input material for sequencing libraries using the SMARTer Stranded Total RNA-Seq Kit V2 (Takara Bio USA, Inc.) following manufacturer's ecommendations and index codes were added to attribute sequences to each sample.Sequencing libraries were generated using QIAseq miRNA Library Kit (Qiagen, Frederick, MD) following manufacturer's recommendations and index codes were added to attribute sequences to each sample. Reverse transcription (RT) primers with unique molecular indices (UMIs) were introduced to analyze the quantification of miRNA expressions during cDNA synthesis and PCR amplification. At last, library quality was assessed on the Agilent Bioanalyzer 2100 and qPCR. The clustering of the index-coded samples was performed on acBot Cluster Generation System using TruSeq PE Cluster Kitv3-cBot-HS (Illumina, San Diego, CA, USA) according to the manufacturer's instructions. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and paired-end reads were generated