Journal
ACS SYNTHETIC BIOLOGY
Volume 12, Issue 4, Pages 1358-1363Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acssynbio.2c00560
Keywords
Candida glabrata; modular cloning toolkit; Golden Gate cloning; CRISPR interference
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Candida glabrata is an emerging drug-resistant human pathogen that causes severe infections in immunocompromised individuals and is a valuable biotechnology host. However, there is a lack of well-characterized cloning toolkit for this yeast. In this study, a CgTK was developed by characterizing a yeast molecular cloning toolkit and adding missing components, which can be used for genome-wide library screens.
The yeast Candida glabrata is an emerging, often drug-resistant opportunistic human pathogen that can cause severe systemic infections in immunocompromised individuals. At the same time, it is a valuable biotechnology host that naturally accumulates high levels of pyruvate-a valuable chemical precursor. Tools for the facile engineering of this yeast could greatly accelerate studies on its pathogenicity and its optimization for biotechnology. While a few tools for plasmid-based expression and genome engineering have been developed, there is no well-characterized cloning toolkit that would allow the modular assembly of pathways or genetic circuits. Here, by characterizing the Saccharomyces cerevisiae-based yeast molecular cloning toolkit (YTK) in C. glabrata and by adding missing components, we build a well-characterized CgTK (C. glabrata toolkit). We used the CgTK to build a CRISPR interference system for C. glabrata that can be used to generate selectable phenotypes via single-gRNA targeting such as is required for genome-wide library screens.
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