title: Targeted miRNAseq of Pediatric Thyroid Cancers with DICER1 Rnase IIIb mutation
design: HTG EdgeSeq Whole Transcriptome miRNA Assay (Tucson, Arizona). FFPE tissue sections (5m) were processed by adding HTG Lysis Buffer to lyse and/or permeabilize the RNA available to subsequently bind to corresponding target-specific Nuclease Protection Probes (NPPs). Target capture was done by adding NPPs to the lysed samples to hybridize to the target miRNA. Then S1 nuclease was added to digest non-hybridized miRNA and excess NPPs, leaving target miRNA/NPP duplexes. S1 digestion was terminated by heat denaturation of S1 enzyme. The processed samples were used as template to set up PCR reactions with primer tags. These tags share common sequences that are complementary to 5'-end and 3'-end wing sequences of the probes and common adaptors required for cluster generation for deep sequencing. The library was prepared using HTG EdgeSeq PCR processing. Following library preparation, it was quantified using HTG EdgeSeq KAPA Library Quantification for Illumina Sequencing.