Feb 27, 2020

Multiplexed conditional genome editing with Cas12a in Drosophila

bioRxiv
Fillip PortMichael Boutros

Abstract

CRISPR-Cas genome engineering has revolutionised biomedical research by enabling targeted genome modification with unprecedented ease. In the popular model organism Drosophila melanogaster gene editing has so far relied exclusively on the prototypical CRISPR nuclease Cas9. The availability of additional CRISPR systems could expand the genomic target space, offer additional modes of regulation and enable the independent manipulation of genes in different cell populations of the same animal. Here we describe a platform for efficient Cas12a gene editing in Drosophila. We show that Cas12a from Lachnospiraceae bacterium, but not Acidaminococcus spec., can mediate robust gene editing in vivo. In combination with most crRNAs, LbCas12a activity is strongly suppressed at lower temperatures, enabling control of gene editing by simply modulating temperature. LbCas12a can directly utilize compact crRNAs arrays that are substantially easier to construct than Cas9 sgRNA arrays, facilitating multiplex genome engineering of several target sites in parallel. Targeting genes with arrays of three crRNAs results in the induction of loss-of function phenotypes with comparable efficiencies than a state-of-the-art Cas9 system. Lastly, we show that ce...Continue Reading

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Mentioned in this Paper

Analysis
Expanding
RNA, Guide
Genetic Engineering
CDC42SE1
Finding
Clustered Regularly Interspaced Short Palindromic Repeats
Drosophila
Protein Expression
Regulation of Biological Process

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