biostudies-literatureHeng EAgency for Science, Technology and ResearchNational Research Foundation SingaporeInstitute of Molecular and Cell Biology253-261https://www.ebi.ac.uk/biostudies/studies/S-EPMC10060144biostudies-literatureUnknown8(2)With the advent of rapid automated <i>in silico</i> identification of biosynthetic gene clusters (BGCs), genomics presents vast opportunities to accelerate natural product (NP) discovery. However, prolific NP producers, <i>Streptomyces</i>, are exceptionally GC-rich (>80%) and highly repetitive within BGCs. These pose challenges in sequencing and high-quality genome assembly which are currently circumvented <i>via</i> intensive sequencing. Here, we outline a more cost-effective workflow using multiplex Illumina and Oxford Nanopore sequencing with hybrid long-short read assembly algorithms to generate high quality genomes. Our protocol involves subjecting long read-derived assemblies to up to 4 rounds of polishing with short reads to yield accurate BGC predictions. We successfully sequencedSynthetic and systems biotechnologyCost-effective hybrid long-short read assembly delineates alternative GC-rich <i>Streptomyces</i> hosts for natural product discovery.PMC1006014421719NRF-CRP19-2017-05-00Yang LKKanagasundaram YLeong CYKoduru LTay DWPSeow DCSHeng ELim YHWong FTNg VNg SBTan LLfalseCost-effective hybrid long-short read assembly delineates alternative GC-rich <i>Streptomyces</i> hosts for natural product discovery.With the advent of rapid automated <i>in silico</i> identification of biosynthetic gene clusters (BGCs), genomics presents vast opportunities to accelerate natural product (NP) discovery. However, prolific NP producers, <i>Streptomyces</i>, are exceptionally GC-rich (>80%) and highly repetitive within BGCs. These pose challenges in sequencing and high-quality genome assembly which are currently circumvented <i>via</i> intensive sequencing. Here, we outline a more cost-effective workflow using multiplex Illumina and Oxford Nanopore sequencing with hybrid long-short read assembly algorithms to generate high quality genomes. Our protocol involves subjecting long read-derived assemblies to up to 4 rounds of polishing with short reads to yield accurate BGC predictions. We successfully sequenced2023-01-01T00:00:00Z2023 Jun2026-07-15T01:27:11.334Z2025-02-19T02:23:02.243ZS-EPMC100601443700727710.1016/j.synbio.2023.03.001