GSE157916
Small RNA sequencing of cerebrospinal fluid exosome fraction in medulloblastoma patients
We reported that the microRNA profiling of exosome isolated from 58 cerebrospinal fluid (CSF) of 38 medulloblastoma (MBL) patient with or without letomeningeal metastases (LM). CSF exosome was precipitated using Exo2DTM for RNA assay (EXOSOME plus, Suwon, South Korea) according to the manufacture's protocol (Kim, et al. 2017). 2 ml of CSF were added to Exo2DTM reagent B following mixing by inverting, and incubated at room temperature for 30 min. The supernatant was discarded and the exosome containing pellet was resuspended in 100 ul of PBS to further RNA isolation. Total RNA was extracted from the CSF exosome fraction using a microRNA purification kit (Norgen Biotek corp., Thorold, Canada) according to the manufacture's protocol. Briefly, samples were mixed with lysis buffer, and the lysate was added to absolute ethanol. The lysate was applied up to the column and centrifuged at 8,000 rpm for 1 min. The column was washed with wash solution and centrifuged at 14,000 rpm for 1 min. Finally, the column was applied to 20 ul of elution solution and centrifuged at 14,000 rpm for 2 min, and the flow-through was stored at –80 ? for storage. RNA precipitates were quantified and qualified with Bioanalyzer Pico 6000 Chip (Agilent Technologies, Santa Clara, CA). The 10 ng of total RNA was provided to the cDNA library generation using SMARTer smRNA-Seq kit (Illumina Inc., San Diego, CA), according to the user manual. The cDNA library was subjected to sequencing using Truseq SBS Kit and HiSeq 2500 System (Illumina Inc., San Diego, CA), according to user guide. Overall design: 58 cerebrospinal fluid (CSF) samples of medulloblastoma patients