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  1. Article ; Online: Author Correction: Inducible expression of large gRNA arrays for multiplexed CRISPRai applications.

    Shaw, William M / Studená, Lucie / Roy, Kyler / Hapeta, Piotr / McCarty, Nicholas S / Graham, Alicia E / Ellis, Tom / Ledesma-Amaro, Rodrigo

    Nature communications

    2023  Volume 14, Issue 1, Page(s) 137

    Language English
    Publishing date 2023-01-10
    Publishing country England
    Document type Published Erratum
    ZDB-ID 2553671-0
    ISSN 2041-1723 ; 2041-1723
    ISSN (online) 2041-1723
    ISSN 2041-1723
    DOI 10.1038/s41467-023-35867-9
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: Inducible expression of large gRNA arrays for multiplexed CRISPRai applications.

    Shaw, William M / Studená, Lucie / Roy, Kyler / Hapeta, Piotr / McCarty, Nicholas S / Graham, Alicia E / Ellis, Tom / Ledesma-Amaro, Rodrigo

    Nature communications

    2022  Volume 13, Issue 1, Page(s) 4984

    Abstract: CRISPR gene activation and inhibition (CRISPRai) has become a powerful synthetic tool for influencing the expression of native genes for foundational studies, cellular reprograming, and metabolic engineering. Here we develop a method for near leak-free, ... ...

    Abstract CRISPR gene activation and inhibition (CRISPRai) has become a powerful synthetic tool for influencing the expression of native genes for foundational studies, cellular reprograming, and metabolic engineering. Here we develop a method for near leak-free, inducible expression of a polycistronic array containing up to 24 gRNAs from two orthogonal CRISPR/Cas systems to increase CRISPRai multiplexing capacity and target gene flexibility. To achieve strong inducibility, we create a technology to silence gRNA expression within the array in the absence of the inducer, since we found that long gRNA arrays for CRISPRai can express themselves even without promoter. Using this method, we create a highly tuned and easy-to-use CRISPRai toolkit in the industrially relevant yeast, Saccharomyces cerevisiae, establishing the first system to combine simultaneous activation and repression, large multiplexing capacity, and inducibility. We demonstrate this toolkit by targeting 11 genes in central metabolism in a single transformation, achieving a 45-fold increase in succinic acid, which could be precisely controlled in an inducible manner. Our method offers a highly effective way to regulate genes and rewire metabolism in yeast, with principles of gRNA array construction and inducibility that should extend to other chassis organisms.
    MeSH term(s) CRISPR-Cas Systems ; Gene Editing/methods ; RNA, Guide, CRISPR-Cas Systems/genetics ; RNA, Guide, CRISPR-Cas Systems/metabolism ; Saccharomyces cerevisiae/genetics ; Saccharomyces cerevisiae/metabolism ; Transcriptional Activation
    Chemical Substances RNA, Guide, CRISPR-Cas Systems
    Language English
    Publishing date 2022-08-25
    Publishing country England
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2553671-0
    ISSN 2041-1723 ; 2041-1723
    ISSN (online) 2041-1723
    ISSN 2041-1723
    DOI 10.1038/s41467-022-32603-7
    Database MEDical Literature Analysis and Retrieval System OnLINE

    More links

    Kategorien

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

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