Next-generation sequencing (NGS) enables the analysis of both microRNA expression and sequence, allowing for elucidation of a comprehensive landscape of miRNAs in a given tissue and sample type. NGS analysis requires high-quality RNA extraction and preparation of microRNA libraries. In this chapter, we describe the methods used for RNA extraction from tissue specimens, serum, cytological slides, and formalin-fixed paraffin-embedded samples. Although the described library preparation and sequencing approaches are based on Illumina NextSeq 500 sequencing technology, the presented principles shall be compatible with other commercially available sequencing platforms.
Keywords: FFPE samples; NGS; Next-generation sequencing; RNA extraction; Serum samples; microRNA identification; microRNA quantification.