471690 Scaleable Assembly and Characterization of Fungal Natural Product Gene Clusters
Here we describe the realization of a pipeline for the high-throughput characterization of fungal NP biosynthetic gene clusters. Our pipeline encompasses:
bioinformatic tools for identification and prioritization of BGCs, and tools for sequence editing (e.g. intron removal) for improved expression in S. cerevisiae
- an assembly platform that combines commercial gene synthesis and yeast homologous recombination for the cost-effective construction of large (> 20 kb) DNAs
- high-throughput untargeted LC-MS based metabolomics screening of recombinant yeast cultures for the rapid detection and prioritization of potential novel compounds
With this pipeline, we successfully detected potentially novel NPs in four out of six previously unstudied fungal BGCs, and have structurally characterized a subset of the BGC-associated compounds. Our pipeline shows how high-throughput synthetic biology tools can facilitate the rapid discovery and biosynthetic production of complex chemical scaffolds of potential pharmaceutical relevance.