GSM5061316: PD7; Homo sapiens; miRNA-Seq
Frozen plasma samples were thawed in thermostat water bath for 2 min at 37 °C and then centrifuged at 2,500 × g for 15 min at 4 °C to remove precipitated proteins and lipids. ~0.5-1 ml plasma was used as the starting material for each EVs isolation by exoRNeasy Midi kit (Qiagen; Cat#77144) according to the manufacturer's instructions. For EV-RNA extraction, 1 ml Trizol reagent (Invitrogen; Cat#15596026) was added to the column and centrifuged at 3,000 × g for 1 min. Total EV-RNA was isolated by Trizol reagent according to manufacturer's recommendations. A total amount of 0.5-1 ng RNA per sample was used as input material for the RNA sample preparations. The cDNA libraries establishment of small RNAs included adaptor ligation, adapter depletion, first-strand cDNA synthesis, Cas9/sgRNA in vitro cleavage treatment, PCR amplification and gel purification.