CRISPR-Cas-Assisted Multiplexing (CAM): Simple Same-Day Multi-Locus Engineering in Yeast

Demands on the industrial and academic yeast strain engineer have increased significantly in the era of synthetic biology. Installing complex biosynthetic pathways and combining point mutations are tedious and time‐consuming using traditional methods. With multiplex engineering tools, these tasks ca...

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Bibliographic Details
Published in:Journal of cellular physiology 2016-12, Vol.231 (12), p.2563-2569
Main Authors: Walter, Jessica M., Chandran, Sunil S., Horwitz, Andrew A.
Format: Article
Language:English
Subjects:
Biosynthetic Pathways - genetics
Cloning
CRISPR-Cas Systems - genetics
Genetic Engineering
Genetic Loci
Reagents
RNA, Guide, CRISPR-Cas Systems - genetics
Saccharomyces cerevisiae - genetics
Yeasts
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Summary:Demands on the industrial and academic yeast strain engineer have increased significantly in the era of synthetic biology. Installing complex biosynthetic pathways and combining point mutations are tedious and time‐consuming using traditional methods. With multiplex engineering tools, these tasks can be completed in a single step, typically achieving up to sixfold compression in strain engineering timelines. To capitalize on this potential, a variety of yeast CRISPR‐Cas methods have been developed, differing largely in how the guide RNA (gRNA) reagents that direct the Cas9 nuclease are delivered. However, in nearly all reported protocols, the time savings of multiplexing is offset by multiple days of cloning to prepare the required reagents. Here, we discuss the advantages and opportunities of CRISPR‐Cas‐assisted multiplexing (CAM), a same‐day, cloning‐free method for multi‐locus engineering in yeast. J. Cell. Physiol. 231: 2563–2569, 2016. © 2016 Wiley Periodicals, Inc. Installing complex biosynthetic pathways and combining point mutations in yeast are tedious and time‐consuming using traditional methods. Here, we discuss the advantages and opportunities of CRISPR‐Cas‐assisted multiplexing (CAM), a same‐day, cloning‐free method for multi‐locus engineering.
ISSN:0021-9541
1097-4652
DOI:10.1002/jcp.25375