Exosomal miRNAs differences analysis between C26 and MC38

MiRNAs contained in C26-derived and MC38-derived exosomes were determined by IlluminaHiSeq platform. High-throughput sequencing was conducted at Shanghai MajorbioBiopharm TechnologyCo., Ltd. (Shanghai, China). In brief, total RNA from exosomes was prepared and quantified with a NanoDrop ND-2000(NanoDrop Technologies).RNA integrity was assessed using a 2100 Bioanalyzer (Agilent Technologies,Santa Clara, CA, USA). A total amount of 3 µg total RNA per sample was used as input material for the small RNA library. Small RNA adapters were then ligated to the 5' and 3' ends of total RNA. After cDNA synthesis and amplification, the PCR-amplified fragments were purified from the PAGE gel, and the completed cDNA libraries were quantified by an Agilent 2100 Bioanalyzer. Cluster generation was performed on an IlluminacBot, and sequencing was performed on an IlluminaHiSeq 2000 platform and 50bp single-end reads were generated.The expression level of each miRNA was calculated according to the transcripts per million reads (TPM) method. Significant differently expressed (DE) miRNAs were extracted with |log2FC|>1 and FDR < 0.05 by DEseq2. Overall design: Exosomes were extracted from conditioned medium of C26 and MC38 cells. MiRNAs contained in C26-derived and MC38-derived exosomes were determined by IlluminaHiSeq platform.