Exorneasy serum plasma maxi kit

Exosomes from human and mouse plasma were extracted using an Invitrogen™ Total Exosome Isolation Kit (ThermoFisher, USA) [27 (link)]. Briefly, 200 uL of plasma was mixed with the precipitation reagent provided by the kit and exosomes were isolated according to the manufacturer’s instructions. The exoRNeasy Serum/Plasma Maxi Kit (Yeasen Biotech, Shanghai, China) [28 (link)] was employed to isolate exosomal RNAs in samples previously spiked with 25 fmol of C. elegans cel-miR-39 standard RNA (Sangon, Shanghai, China), used as control. The exosomes were stored at -80° C before cell culture experiments and molecular analyses.

Total RNA in cells and tissues were obtained using TRIzol reagent. Rayscript cDNA synthesis kit (GENEray Biotechnology, Shanghai, China) was used to perform the cDNA synthesis. After adding SYBR green reagent (GENEray Biotechnology) and primers in cDNA solution, amplification and real‐time fluorescence detection were conducted. GAPDH was set as a loading control to measure the relative levels of target genes. Exosomal RNAs were isolated utilizing the exoRNeasy Serum/Plasma Maxi Kit (Yeasen Biotech, Shanghai, China), which had been pre‐enriched with 25 fmol of C. elegans cel‐miR‐39 standard RNA (Sangon, Shanghai, China). The C. elegans cel‐miR‐39 standard RNA served as a control to standardize the exosomal RNAs. Used primers were shown in Table S1.