Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Development of a Transformation Method for Metschnikowia borealis and Other CUG-Serine Yeasts

Zachary Gordon
,
Maximillian P. M. Soltysiak
ORCID logo ,
Christopher Leichthammer
,
Jasmine A. Therrien
,
Rebecca S. Meaney
,
Carolyn Lauzon
,
Matthew Adams
,
Dong Kyung Lee
,
Preetam Janakirama
,
Marc-Andre Lachance
,
Bogumil J. Karas
* ORCID logo
Version 1 : Received: 10 November 2018 / Approved: 13 November 2018 / Online: 13 November 2018 (05:17:45 CET)

A peer-reviewed article of this Preprint also exists.

Gordon, Z.B.; Soltysiak, M.P.; Leichthammer, C.; Therrien, J.A.; Meaney, R.S.; Lauzon, C.; Adams, M.; Lee, D.K.; Janakirama, P.; Lachance, M.-A.; Karas, B.J. Development of a Transformation Method for Metschnikowia borealis and other CUG-Serine Yeasts. Genes 2019, 10, 78. Gordon, Z.B.; Soltysiak, M.P.; Leichthammer, C.; Therrien, J.A.; Meaney, R.S.; Lauzon, C.; Adams, M.; Lee, D.K.; Janakirama, P.; Lachance, M.-A.; Karas, B.J. Development of a Transformation Method for Metschnikowia borealis and other CUG-Serine Yeasts. Genes 2019, 10, 78.

Abstract

Yeasts belonging to the Metschnikowia genus are particularly interesting for the unusual formation of only two needle-shaped ascospores during their mating cycle. Presently, the meiotic process that can lead to only two spores from a diploid zygote is poorly understood. The expression of fluorescent nuclear proteins should allow the meiotic process to be visualized in vivo; however, no large-spored species of Metschnikowia has ever been transformed. Accordingly, we aimed to develop a transformation method for Metschnikowia borealis, a particularly large-spored species of Metschnikowia, with the goal of enabling the genetic manipulations required to study biological processes in detail. Genetic analyses confirmed that M. borealis, and many other Metchnikowiacea, are CUG-Ser yeasts. Codon-optimized selectable markers lacking CUG codons were used to successfully transform M. borealis by electroporation and lithium acetate, and transformants appeared to be the result of random integration. Mating experiments confirmed that transformed-strains were capable of generating large asci and undergoing recombination. Finally, random integration was used to transform an additional 18 yeast strains, and all attempts successfully generated transformants. The results provide a simple method to transform many yeasts from an array of different clades and can be used to study or develop many species for various applications.

Keywords

genome engineering; synthetic biology; yeasts; Metschnikowia; genetic tools; DNA delivery; CUG-Ser

Subject

Biology and Life Sciences, Biology and Biotechnology

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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