GSM8690989: shLUC_6; Homo sapiens; miRNA-Seq
Approximately 50 mg of fresh-frozen xenograft PT tissue was crushed in liquid nitrogen in the presence of QIAzol Lysis Reagent (Qiagen, Hilden, Germany). The total RNA was isolated using miRNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. All RNA samples were treated with DNase I during the isolation procedure. The RNA yield was determined by UV absorbance using a NanoDrop 1000 spectrophotometer (Peqlab, Erlangen, Germany). The RNA quality was assessed by analyzing the ribosomal RNA integrity number (RIN) on an Agilent 2100 Bioanalyzer using the RNA 6000 Nano kit (Agilent Technologies, Palo Alto, CA, USA). The RIN values of the isolated RNA samples ranged from 6.7 to 9.6. Libraries for miRNA sequencing were prepared from total RNA samples using NEBNext Multiplex Small RNA Library Prep Kit for Illumina. Each sample was sequenced on the Illumina NextSeq 550 to generate single-end 50-nucleotide reads: 7 control samples (shLUC) and 8 samples with CD44 knockdown (shCD44).