| Title |
Genome editing in plants using the compact editor CasΦ / |
| Authors |
Li, Zheng ; Zhong, Zhenhui ; Wu, Zhongshou ; Pausch, Patrick ; Al-Shayeb, Basem ; Amerasekera, Jasmine ; Doudna, Jennifer A ; Jacobsen, Steven E |
| DOI |
10.1073/pnas.2216822120 |
| Full Text |
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| Is Part of |
PNAS(Proceedings of the National Academy of Sciences of the United States of America).. Washington : Published by PNAS. 2023, vol. 120, no. 4, art. no. e2216822120, p. [1-10].. ISSN 0027-8424. eISSN 1091-6490 |
| Keywords [eng] |
CRISPR-CasΦ ; lant genome editing ; DNA methylation ; CasΦ variants ; off-target editing |
| Abstract [eng] |
Clustered regularly interspaced short palindromic repeats and CRISPR-associated pro-teins (CRISPR-Cas) systems have been developed as important tools for plant genome engineering. Here, we demonstrate that the hypercompact CasΦ nuclease is able to generate stably inherited gene edits in Arabidopsis, and that CasΦ guide RNAs can be expressed with either the Pol-III U6 promoter or a Pol-II promoter together with ribozyme mediated RNA processing. Using the Arabidopsis fwa epiallele, we show that CasΦ displays higher editing efficiency when the target locus is not DNA methylated, suggesting that CasΦ is sensitive to chromatin environment. Importantly, two CasΦprotein variants, vCasΦ and nCasΦ, both showed much higher editing efficiency relative to the wild-type CasΦ enzyme. Consistently, vCasΦ and nCasΦ yielded offspring plants with inherited edits at much higher rates compared to WTCasΦ. Extensive genomic analysis of gene edited plants showed no off-target editing, suggesting that CasΦ is highly specific. The hypercompact size, T-rich minimal protospacer adjacent motif (PAM), and wide range of working temperatures make CasΦ an excellent supplement to existing plant genome editing systems. |
| Published |
Washington : Published by PNAS |
| Type |
Journal article |
| Language |
English |
| Publication date |
2023 |
| CC license |
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