Multiplexed CRISPR/Cas9 Genome Editing and Gene Regulation Using Csy4 in Saccharomyces cerevisiae
Artikel i vetenskaplig tidskrift, 2018

Clustered regularly interspaced short palindromic repeats (CRISPR) technology has greatly accelerated the field of strain engineering. However, insufficient efforts have been made toward developing robust multiplexing tools in Saccharomyces cerevisiae. Here, we exploit the RNA processing capacity of the bacterial endoribonuclease Csy4 from Pseudomonas aeruginosa, to generate multiple gRNAs from a single transcript for genome editing and gene interference applications in S. cerevisiae. In regards to genome editing, we performed a quadruple deletion of FAA1, FAA4, POX1 and TES1 reaching 96% efficiency out of 24 colonies tested. Then, we used this system to efficiently transcriptionally regulate the three genes, OLE1, HMG1 and ACS1. Thus, we demonstrate that multiplexed genome editing and gene regulation can be performed in a fast and effective manner using Csy4.

CRISPR

metabolic engineering

multiplexing

gene editing

CRISPRi

Csy4

gene regulation

Författare

Raphael Ferreira

Chalmers, Biologi och bioteknik, Systembiologi

Christos Skrekas

Chalmers, Biologi och bioteknik, Systembiologi

Jens B Nielsen

Danmarks Tekniske Universitet (DTU)

Chalmers, Biologi och bioteknik, Systembiologi

Florian David

Chalmers, Biologi och bioteknik, Systembiologi

ACS Synthetic Biology

2161-5063 (eISSN)

Vol. 7 1 10-15

Ämneskategorier

Mikrobiologi

Bioinformatik och systembiologi

Genetik

DOI

10.1021/acssynbio.7b00259