GSM5035443: Plasma MVs - C26 rep2; Mus musculus; ncRNA-Seq
Serum-free conditioned media or plasma were centrifuged at 300 × g for 10 min to eliminate cell debris. The supernatant was then recovered and successive centrifugations at increasing speed were performed: one at 2,000 × g for 20 min to eliminate dead cells, then one at 10,000 × g for 30 min to remove cell debris, and finally one at 100,000 × g for 70 min to pellet MVs. The resulting pellet was then washed in sterilized PBS and ultracentrifuged again at the same speed. The obtained pellet was finally re-suspended in 1 mL TriPure and total RNA was isolated following the manufacturer's instructions NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England BioLabs Inc., Ipswich, MA, USA);Protocol E7330) The obtained sequence libraries were subjected to the Illumina sequencing pipeline, passing through clonal cluster generation on a single-read flow cell (Illumina Inc., San Diego, CA, USA) by bridge amplification and were sequenced at ultrahigh throughput on NextSeq500 (Illumina Inc., San Diego, CA, USA). One flowcell in 24-plex could produce about six million single reads per sample, 75 base pairs (bp) long
GEO Accession
GSM5035443