Abstract
Genomics has revolutionized the research on fungal secondary metabolite (SM) biosynthesis. To elucidate the molecular and enzymatic mechanisms underlying the biosynthesis of a specific SM compound, the important first step is often to find the genes that responsible for its synthesis. The accessibility to fungal genome sequences allows the bypass of the cumbersome traditional library construction and screening approach. The advance in next-generation sequencing (NGS) technologies have further improved the speed and reduced the cost of microbial genome sequencing in the past few years, which has accelerated the research in this field. Here, we will present an example work flow for identifying the gene cluster encoding the biosynthesis of SMs of interest using an NGS approach. We will also review the different strategies that can be employed to pinpoint the targeted gene clusters rapidly by giving several examples stemming from our work.
Highlights
Human health has been benefited from the secondary metabolites (SMs) produced by fungi
This approach has facilitated the identification of genes for biosynthesis of tryptoquivaline in A. clavatus, which was separated into three genomic loci, by comparing with the tryptoquialanine-producing P. aethiopicum genome (Gao et al, 2011)
Next-generation sequencing technologies have significantly accelerated the process of targeted SM gene cluster discovery
Summary
Human health has been benefited from the secondary metabolites (SMs) produced by fungi. WGS of G. zeae with the Sanger sequencing method has allowed the systematic deletion of all 15 PKS genes in the fungal genome, which lead to identification of the gene cluster for zearalenone, aurofusarin, fusarin C and an unidentified black perithecial pigment (Gaffoor et al, 2005) This opens up the opportunities for detailed characterization of these pathways for zearalenone (Kim et al, 2005; Zhou et al, 2010; Lee et al, 2011), aurofusarin (Frandsen et al, 2006; Frandsen et al, 2011), and fusarin C (Niehaus et al, 2013). The development of next-generation sequencing (NGS) technologies in the last decade has dramatically lowered the cost for DNA sequencing and put the power of microbial WGS in the hand of individual laboratories (van Dijk et al, 2014) This technology revolution has energized the natural product research field and sparked some exciting NGS-based targeted SM gene cluster discovery projects in fungi (Table 1). Our work that used such NGS approach includes the discovery of SM clusters encoding viridicatumtoxin and griseofulvin (Chooi et al, 2010), tryptoquialanine (Gao et al, 2011), echinocandin (Cacho et al, 2012),
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