Transposon mutagenesis in bacterial natural product discovery

Transposon mutagenesis has re-emerged as a powerful and versatile strategy for discovering and characterising specialised metabolites encoded by biosynthetic gene clusters (BGCs). While genomics has revealed an enormous diversity of putative BGCs across bacteria, many remain silent, weakly expressed, or genetically intractable, necessitating experimental tools that can link genotype to chemical output. Transposons provide an unbiased and broadly applicable platform for disrupting, activating, or modulating gene expression without relying on homologous recombination, making them particularly valuable in challenging microbial hosts. Here, we review the major applications of transposon mutagenesis in natural product discovery, providing examples that highlight discoveries made using phenotype- and bioactivity-guided screens, phenotype-independent strategies, and transposon-based engineering of heterologous expression platforms. Transposon technologies provide flexible and scalable tools for activating, characterising, and engineering microbial BGCs. As genome mining continues to unearth rich seams of unexplored metabolic potential, these tools will remain essential for converting genetic predictions into chemical discovery.