Improved betulinic acid biosynthesis using synthetic yeast chromosome recombination and semi-automated rapid LC-MS screening
File(s)
Published version
Author(s)
Type
Journal Article
Abstract
Synthetic biology, genome engineering and directed evolution offer innumerable tools to expedite engineering of strains for optimising biosynthetic pathways. One of the most radical is SCRaMbLE, a system of inducible in vivo deletion and rearrangement of synthetic yeast chromosomes, diversifying the genotype of millions of Saccharomyces cerevisiae cells in hours. SCRaMbLE can yield strains with improved biosynthetic phenotypes but is limited by screening capabilities. To address this bottleneck, we combine automated sample preparation, an ultra-fast 84-second LC-MS method, and barcoded nanopore sequencing to rapidly isolate and characterise the best performing strains. Here, we use SCRaMbLE to optimise yeast strains engineered to produce the triterpenoid betulinic acid. Our semi-automated workflow screens 1,000 colonies, identifying and sequencing 12 strains with between 2- to 7-fold improvement in betulinic acid titre. The broad applicability of this workflow to rapidly isolate improved strains from a variant library makes this a valuable tool for biotechnology.
Date Issued
2020-02-13
Date Acceptance
2020-01-24
Citation
Nature Communications, 2020, 11
ISSN
2041-1723
Publisher
Nature Research (part of Springer Nature)
Journal / Book Title
Nature Communications
Volume
11
Copyright Statement
© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
License URL
Sponsor
Biotechnology and Biological Sciences Research Council (BBSRC)
Biotechnology and Biological Sciences Research Council (BBSRC)
Biotechnology and Biological Sciences Research Council (BBSRC)
Engineering & Physical Science Research Council (EPSRC)
Grant Number
BB/K006290/1
BB/K019791/1
BB/L027852/1
EP/N023854/1
Publication Status
Published
Article Number
ARTN 868
Date Publish Online
2020-02-13