Victoria Link Limited

Massey University

Victoria Link Llimited

Craig John Van Dolleweerd

Emily Jane Parker

Matthew Joseph Nicholson

Leyla Yolanda Bustamante Rodriguez

Sarah Adeline Kessans

David Barry Scott

Kyle Cornelius Van De Bittner

Nodulisporic acids (NAs) comprise a group of indole diterpenes known for their potent insecticidal activities; however, biosynthesis of NAs by its natural producer,(sp.) is exceptionally difficult to achieve. The identification of genes responsible for NA production could enable biosynthetic pathway optimization to provide access to NAs for commercial applications. Obtaining useful quantities of NAs using published fermentations methods is challenging, making gene knockout studies an undesirable method to confirm gene function. Alternatively, heterologous gene expression ofgenes in a more robust host species likeprovides a way to rapidly identify the function of genes that play a role in NA biosynthesis. In this work, we identified the function of four secondary-metabolic genes necessary for the biosynthesis of nodulisporic acid F (NAF) and reconstituted these genes in the genome ofto enable heterologous production of NAF in this fungus.

Heterologous biosynthesis of nodulisporic acid

A modular and hierarchical DNA assembly platform for synthetic biology is described. This enabling technology, termed MIDAS (for Modular Idempotent DNA Assembly System), can precisely assemble multiple DNA fragments in a single reaction using a standardised assembly design. It can be used to build genes from libraries of sequence-verified, reusable parts and to assemble multiple genes in a single vector. We describe the design and use of MIDAS, and its application in the reconstruction of the metabolic pathway for production of paspaline, a key intermediate in the biosynthesis of a range of indole diterpenes&#x2014;a class of economically important secondary metabolites produced by several species of filamentous fungi.

Modular DNA assembly system

Growing community of inventors

Prebbleton, New Zealand

Craig John Van Dolleweerd