Efficient engineering of marker-free synthetic allotetraploids of Saccharomyces

William G. Alexander; David Peris; Brandon T. Pfannenstiel; Dana A. Opulente; Meihua Kuang; Chris Todd Hittinger

doi:10.1016/j.fgb.2015.11.002

Efficient engineering of marker-free synthetic allotetraploids of Saccharomyces

William G. Alexander
, David Peris
, Brandon T. Pfannenstiel
, Dana A. Opulente
, Meihua Kuang
, Chris Todd Hittinger

Research output: Contribution to journal › Article › peer-review

41 Scopus citations

Abstract

Saccharomyces interspecies hybrids are critical biocatalysts in the fermented beverage industry, including in the production of lager beers, Belgian ales, ciders, and cold-fermented wines. Current methods for making synthetic interspecies hybrids are cumbersome and/or require genome modifications. We have developed a simple, robust, and efficient method for generating allotetraploid strains of prototrophic Saccharomyces without sporulation or nuclear genome manipulation. S. cerevisiae × S. eubayanus, S. cerevisiae × S. kudriavzevii, and S. cerevisiae × S. uvarum designer hybrid strains were created as synthetic lager, Belgian, and cider strains, respectively. The ploidy and hybrid nature of the strains were confirmed using flow cytometry and PCR-RFLP analysis, respectively. This method provides an efficient means for producing novel synthetic hybrids for beverage and biofuel production, as well as for constructing tetraploids to be used for basic research in evolutionary genetics and genome stability.

Original language	English
Pages (from-to)	10-17
Number of pages	8
Journal	Fungal Genetics and Biology
Volume	89
DOIs	https://doi.org/10.1016/j.fgb.2015.11.002
State	Published - Apr 1 2016
Externally published	Yes

Funding

We would like to thank James Hose and Audrey Gasch for the training with and use of their Guava easyCyte flow cytometer; Carol Newlon and Lucia Fabiani for the KARS101 plasmid; and Trey K. Sato for the NRRL YB-210 strain. This material is based upon work supported by the National Science Foundation under Grant Nos. DEB-1253634 and DEB-1442148 to CTH and funded in part by the DOE Great Lakes Bioenergy Research Center (DOE Office of Science BER DE-FC02-07ER64494 ). CTH is an Alfred Toepfer Faculty Fellow, supported by the Alexander von Humboldt Foundation . CTH is a Pew Scholar in the Biomedical Sciences, supported by the Pew Charitable Trusts . BP was supported by the Predoctoral Training Program in Genetics, funded by the National Institutes of Health ( 5 T32 GM007133-40 ).

Keywords

Biofuels
Brewing
Hybrids
Prototrophic
Saccharomyces
Synthetic zymurgy

Access to Document

10.1016/j.fgb.2015.11.002

Cite this

@article{aae095b9958b4bfd9db5e83261e4066f,

title = "Efficient engineering of marker-free synthetic allotetraploids of Saccharomyces",

abstract = "Saccharomyces interspecies hybrids are critical biocatalysts in the fermented beverage industry, including in the production of lager beers, Belgian ales, ciders, and cold-fermented wines. Current methods for making synthetic interspecies hybrids are cumbersome and/or require genome modifications. We have developed a simple, robust, and efficient method for generating allotetraploid strains of prototrophic Saccharomyces without sporulation or nuclear genome manipulation. S. cerevisiae × S. eubayanus, S. cerevisiae × S. kudriavzevii, and S. cerevisiae × S. uvarum designer hybrid strains were created as synthetic lager, Belgian, and cider strains, respectively. The ploidy and hybrid nature of the strains were confirmed using flow cytometry and PCR-RFLP analysis, respectively. This method provides an efficient means for producing novel synthetic hybrids for beverage and biofuel production, as well as for constructing tetraploids to be used for basic research in evolutionary genetics and genome stability.",

keywords = "Biofuels, Brewing, Hybrids, Prototrophic, Saccharomyces, Synthetic zymurgy",

author = "Alexander, \{William G.\} and David Peris and Pfannenstiel, \{Brandon T.\} and Opulente, \{Dana A.\} and Meihua Kuang and Hittinger, \{Chris Todd\}",

note = "Publisher Copyright: {\textcopyright} 2015 Elsevier Inc.",

year = "2016",

month = apr,

day = "1",

doi = "10.1016/j.fgb.2015.11.002",

language = "English",

volume = "89",

pages = "10--17",

journal = "Fungal Genetics and Biology",

issn = "1087-1845",

publisher = "Elsevier",

}

TY - JOUR

T1 - Efficient engineering of marker-free synthetic allotetraploids of Saccharomyces

AU - Alexander, William G.

AU - Peris, David

AU - Pfannenstiel, Brandon T.

AU - Opulente, Dana A.

AU - Kuang, Meihua

AU - Hittinger, Chris Todd

PY - 2016/4/1

Y1 - 2016/4/1

N2 - Saccharomyces interspecies hybrids are critical biocatalysts in the fermented beverage industry, including in the production of lager beers, Belgian ales, ciders, and cold-fermented wines. Current methods for making synthetic interspecies hybrids are cumbersome and/or require genome modifications. We have developed a simple, robust, and efficient method for generating allotetraploid strains of prototrophic Saccharomyces without sporulation or nuclear genome manipulation. S. cerevisiae × S. eubayanus, S. cerevisiae × S. kudriavzevii, and S. cerevisiae × S. uvarum designer hybrid strains were created as synthetic lager, Belgian, and cider strains, respectively. The ploidy and hybrid nature of the strains were confirmed using flow cytometry and PCR-RFLP analysis, respectively. This method provides an efficient means for producing novel synthetic hybrids for beverage and biofuel production, as well as for constructing tetraploids to be used for basic research in evolutionary genetics and genome stability.

AB - Saccharomyces interspecies hybrids are critical biocatalysts in the fermented beverage industry, including in the production of lager beers, Belgian ales, ciders, and cold-fermented wines. Current methods for making synthetic interspecies hybrids are cumbersome and/or require genome modifications. We have developed a simple, robust, and efficient method for generating allotetraploid strains of prototrophic Saccharomyces without sporulation or nuclear genome manipulation. S. cerevisiae × S. eubayanus, S. cerevisiae × S. kudriavzevii, and S. cerevisiae × S. uvarum designer hybrid strains were created as synthetic lager, Belgian, and cider strains, respectively. The ploidy and hybrid nature of the strains were confirmed using flow cytometry and PCR-RFLP analysis, respectively. This method provides an efficient means for producing novel synthetic hybrids for beverage and biofuel production, as well as for constructing tetraploids to be used for basic research in evolutionary genetics and genome stability.

KW - Biofuels

KW - Brewing

KW - Hybrids

KW - Prototrophic

KW - Saccharomyces

KW - Synthetic zymurgy

UR - https://www.scopus.com/pages/publications/84960110043

U2 - 10.1016/j.fgb.2015.11.002

DO - 10.1016/j.fgb.2015.11.002

M3 - Article

C2 - 26555931

AN - SCOPUS:84960110043

SN - 1087-1845

VL - 89

SP - 10

EP - 17

JO - Fungal Genetics and Biology

JF - Fungal Genetics and Biology

ER -

Efficient engineering of marker-free synthetic allotetraploids of Saccharomyces

Abstract

Funding

Keywords

Access to Document

Other files and links

Fingerprint

Cite this