MicroRNAs are important regulators of gene expression that control both physiological and pathological processes such as development and cancer. Although their mode of action has attracted great attention, the principles governing their expression and activity are only beginning to emerge. Recent studies have introduced a paradigm shift in our understanding of the microRNA biogenesis pathway, which was previously believed to be universal to all microRNAs. Maturation steps specific to individual microRNAs have been uncovered, and these offer a plethora of regulatory options after transcription with multiple proteins affecting microRNA processing efficiency. Here we review the recent advances in knowledge of the microRNA biosynthesis pathways and discuss their impact on post-transcriptional microRNA regulation during tumour development.