Extraction and Oxford Nanopore sequencing of genomic DNA from filamentous Actinobacteria

Actinomycetota (Actinobacteria) is an ecologically and industrially important phylum which is challenging to extract pure high-molecular-weight (HMW) DNA from. This protocol provides a parallelized, cost-effective, and straightforward approach for consistently extracting pure HMW DNA using modified non-toxic commercial kits suitable for higher throughput applications. We further provide a workflow for sequencing and assembly of complete genomes using an optimized Oxford Nanopore rapid barcoding protocol and Illumina data error correction.