Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides
Abstract
The oleaginous yeast Rhodosporidium toruloides is considered a promising candidate for production of chemicals and biofuels thanks to its ability to grow on lignocellulosic biomass, and its high production of lipids and carotenoids. However, efforts to engineer this organism are hindered by a lack of suitable genetic tools. In this paper, we report the development of a CRISPR/Cas9 system for genome editing in R. toruloides based on a fusion 5S rRNA–tRNA promoter for guide RNA (gRNA) expression, capable of greater than 95% gene knockout for various genetic targets. Additionally, multiplexed double-gene knockout mutants were obtained using this method with an efficiency of 78%. Finally, this tool can be used to accelerate future metabolic engineering work in this yeast.
- Authors:
-
- Univ. of Illinois at Urbana‐Champaign, Urbana, IL (United States)
- Publication Date:
- Research Org.:
- Center for Advanced Bioenergy and Bioproducts Innovation (CABBI), Urbana, IL (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1542927
- Alternate Identifier(s):
- OSTI ID: 1515089
- Grant/Contract Number:
- SC0018420; SC0018260
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Biotechnology and Bioengineering
- Additional Journal Information:
- Journal Volume: 116; Journal Issue: 8; Journal ID: ISSN 0006-3592
- Publisher:
- Wiley
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; 09 BIOMASS FUELS; CRISPR/Cas9; genome editing; metabolic engineering; multiplex; Rhodosporidium toruloides
Citation Formats
Schultz, J. Carl, Cao, Mingfeng, and Zhao, Huimin. Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides. United States: N. p., 2019.
Web. doi:10.1002/bit.27001.
Schultz, J. Carl, Cao, Mingfeng, & Zhao, Huimin. Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides. United States. https://doi.org/10.1002/bit.27001
Schultz, J. Carl, Cao, Mingfeng, and Zhao, Huimin. Tue .
"Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides". United States. https://doi.org/10.1002/bit.27001. https://www.osti.gov/servlets/purl/1542927.
@article{osti_1542927,
title = {Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides},
author = {Schultz, J. Carl and Cao, Mingfeng and Zhao, Huimin},
abstractNote = {The oleaginous yeast Rhodosporidium toruloides is considered a promising candidate for production of chemicals and biofuels thanks to its ability to grow on lignocellulosic biomass, and its high production of lipids and carotenoids. However, efforts to engineer this organism are hindered by a lack of suitable genetic tools. In this paper, we report the development of a CRISPR/Cas9 system for genome editing in R. toruloides based on a fusion 5S rRNA–tRNA promoter for guide RNA (gRNA) expression, capable of greater than 95% gene knockout for various genetic targets. Additionally, multiplexed double-gene knockout mutants were obtained using this method with an efficiency of 78%. Finally, this tool can be used to accelerate future metabolic engineering work in this yeast.},
doi = {10.1002/bit.27001},
journal = {Biotechnology and Bioengineering},
number = 8,
volume = 116,
place = {United States},
year = {Tue Apr 30 00:00:00 EDT 2019},
month = {Tue Apr 30 00:00:00 EDT 2019}
}
Web of Science
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