GSM8693233: Brain, metastasis, rep1 (miRNA-Seq); Homo sapiens; miRNA-Seq
For total RNA purification, 30 mg/sample of flash-frozen tissue samples were dissected on ice. The tissue samples were disrupted and homogenized with a MagNa Lyser device (Roche Ltd., Switzerland). MiRNA-enriched total RNA isolation was performed according to the manufacturer's instructions using the miRNeasy Mini Kit (Qiagen, Hilden, Germany). The quantity and quality of the isolated RNA were determined using a Nanodrop spectrophotometer (Thermo Scientific, Waltham, MA, USA). The NEBNext Multiplex Small RNA Perp Set for Illumina (1-48) 96 rxn kit (New England BioLabs, Ipswich, MA, USA) was used for library preparation, for which the quality of the RNA samples was examined with the Eukaryotic Total RNA Nano Assay on the Agilent Bio Analyzer (Agilent Technologies, Santa Clara, CA, USA). For small RNA-seq library, 1 µg of total RNA with an RNA integrity number (RIN) score >7 was used. Fragment size distribution and molarity of libraries were also checked on Agilent BioAnalyzer DNA1000 chip (Agilent Technologies, Santa Clara, CA, USA). Finally, the libraries were sequenced on an Illumina NextSeq 500 Sequencing System (Illumina, San Diego, CA, USA) with read lengths of 50 base pairs (single reads). Fastq files were aligned to the human reference genome (GRCh38) using the Novoalign algorithm.