SWITCH: a dynamic CRISPR tool for genome engineering and metabolic pathway control for cell factory construction in Saccharomyces cerevisiae

Katherina Garcia Vanegas, Beata Joanna Lehka, Uffe Hasbro Mortensen

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

Resumé

Background

The yeast Saccharomyces cerevisiae is increasingly used as a cell factory. However, cell factory construction time is a major obstacle towards using yeast for bio-production. Hence, tools to speed up cell factory construction are desirable.


Results

In this study, we have developed a new Cas9/dCas9 based system, SWITCH, which allows Saccharomyces cerevisiae strains to iteratively alternate between a genetic engineering state and a pathway control state. Since Cas9 induced recombination events are crucial for SWITCH efficiency, we first developed a technique TAPE, which we have successfully used to address protospacer efficiency. As proof of concept of the use of SWITCH in cell factory construction, we have exploited the genetic engineering state of a SWITCH strain to insert the five genes necessary for naringenin production. Next, the naringenin cell factory was switched to the pathway control state where production was optimized by downregulating an essential gene TSC13, hence, reducing formation of a byproduct.


Conclusions

We have successfully integrated two CRISPR tools, one for genetic engineering and one for pathway control, into one system and successfully used it for cell factory construction.
OriginalsprogEngelsk
TidsskriftMicrobial Cell Factories
Vol/bind16
Sider (fra-til)25-36
Antal sider12
DOI
StatusUdgivet - 2017

Emneord

  • CRISPR tool
  • genome engineering
  • metabolic pathway control
  • cell factory
  • Saccharomyces cerevisiae

Citer dette

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title = "SWITCH: a dynamic CRISPR tool for genome engineering and metabolic pathway control for cell factory construction in Saccharomyces cerevisiae",
abstract = "BackgroundThe yeast Saccharomyces cerevisiae is increasingly used as a cell factory. However, cell factory construction time is a major obstacle towards using yeast for bio-production. Hence, tools to speed up cell factory construction are desirable.ResultsIn this study, we have developed a new Cas9/dCas9 based system, SWITCH, which allows Saccharomyces cerevisiae strains to iteratively alternate between a genetic engineering state and a pathway control state. Since Cas9 induced recombination events are crucial for SWITCH efficiency, we first developed a technique TAPE, which we have successfully used to address protospacer efficiency. As proof of concept of the use of SWITCH in cell factory construction, we have exploited the genetic engineering state of a SWITCH strain to insert the five genes necessary for naringenin production. Next, the naringenin cell factory was switched to the pathway control state where production was optimized by downregulating an essential gene TSC13, hence, reducing formation of a byproduct.ConclusionsWe have successfully integrated two CRISPR tools, one for genetic engineering and one for pathway control, into one system and successfully used it for cell factory construction.",
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SWITCH: a dynamic CRISPR tool for genome engineering and metabolic pathway control for cell factory construction in Saccharomyces cerevisiae. / Garcia Vanegas, Katherina; Lehka, Beata Joanna; Mortensen, Uffe Hasbro.

I: Microbial Cell Factories, Bind 16, 2017, s. 25-36.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

TY - JOUR

T1 - SWITCH: a dynamic CRISPR tool for genome engineering and metabolic pathway control for cell factory construction in Saccharomyces cerevisiae

AU - Garcia Vanegas, Katherina

AU - Lehka, Beata Joanna

AU - Mortensen, Uffe Hasbro

N1 - The Erratum to this article has been published in Microbial Cell Factories 2017 16:53

PY - 2017

Y1 - 2017

N2 - BackgroundThe yeast Saccharomyces cerevisiae is increasingly used as a cell factory. However, cell factory construction time is a major obstacle towards using yeast for bio-production. Hence, tools to speed up cell factory construction are desirable.ResultsIn this study, we have developed a new Cas9/dCas9 based system, SWITCH, which allows Saccharomyces cerevisiae strains to iteratively alternate between a genetic engineering state and a pathway control state. Since Cas9 induced recombination events are crucial for SWITCH efficiency, we first developed a technique TAPE, which we have successfully used to address protospacer efficiency. As proof of concept of the use of SWITCH in cell factory construction, we have exploited the genetic engineering state of a SWITCH strain to insert the five genes necessary for naringenin production. Next, the naringenin cell factory was switched to the pathway control state where production was optimized by downregulating an essential gene TSC13, hence, reducing formation of a byproduct.ConclusionsWe have successfully integrated two CRISPR tools, one for genetic engineering and one for pathway control, into one system and successfully used it for cell factory construction.

AB - BackgroundThe yeast Saccharomyces cerevisiae is increasingly used as a cell factory. However, cell factory construction time is a major obstacle towards using yeast for bio-production. Hence, tools to speed up cell factory construction are desirable.ResultsIn this study, we have developed a new Cas9/dCas9 based system, SWITCH, which allows Saccharomyces cerevisiae strains to iteratively alternate between a genetic engineering state and a pathway control state. Since Cas9 induced recombination events are crucial for SWITCH efficiency, we first developed a technique TAPE, which we have successfully used to address protospacer efficiency. As proof of concept of the use of SWITCH in cell factory construction, we have exploited the genetic engineering state of a SWITCH strain to insert the five genes necessary for naringenin production. Next, the naringenin cell factory was switched to the pathway control state where production was optimized by downregulating an essential gene TSC13, hence, reducing formation of a byproduct.ConclusionsWe have successfully integrated two CRISPR tools, one for genetic engineering and one for pathway control, into one system and successfully used it for cell factory construction.

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KW - genome engineering

KW - metabolic pathway control

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KW - Saccharomyces cerevisiae

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KW - genome engineering

KW - metabolic pathway control

KW - cell factory

KW - Saccharomyces cerevisiae

U2 - 10.1186/s12934-017-0632-x

DO - 10.1186/s12934-017-0632-x

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JO - Microbial Cell Factories

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SN - 1475-2859

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