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Live imaging-assisted domain-specific CRISPR genome editing at single cell resolution in plants

Li, Ting and Yan, An and Meyerowitz, Elliot M. (2019) Live imaging-assisted domain-specific CRISPR genome editing at single cell resolution in plants. . (Unpublished) https://resolver.caltech.edu/CaltechAUTHORS:20191007-135703984

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Abstract

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology has been widely used for genome engineering in a wide range of organisms, but much of the development of CRISPR-based genome editing has been aimed toward improving its efficiency and accuracy, so as to obtain genetic materials carrying known and stably heritable genome modifications. Precise spatiotemporal control over genome editing technology at cell type resolution is a key challenge for gene function studies. Some tissue-specific CRISPR genome editing methods relying on phenotypic characterization and fluorescent immune-staining techniques have been developed for biomedical research and gene therapy, they function by spatially controlling expression of Cas9. Recent work establishes the presence and location of mutational events at a single cell level in Arabidopsis roots and stomata. Here we present an efficient domain-specific CRISPR-Cas9 system combined with a high resolution live-imaging based screening strategy, applied in the shoot apical meristem of Arabidopsis thaliana. Using the system we investigate PIN-FORMED1 (PIN1) protein functions in tissue morphogenesis and PIN1 mechanical stress response in a cell layer-specific fashion. We find that reported failure to generate new primordia in epidermal PIN1 knockout SAMs is due to a reduction in mechanical stress differences in the sub-epidermal layer. The methods described are applicable to spatial-temporal gene manipulation in plants.


Item Type:Report or Paper (Discussion Paper)
Related URLs:
URLURL TypeDescription
https://doi.org/10.1101/793240DOIDiscussion Paper
ORCID:
AuthorORCID
Li, Ting0000-0001-8273-5593
Yan, An0000-0002-4676-5442
Meyerowitz, Elliot M.0000-0003-4798-5153
Additional Information:The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission. bioRxiv preprint first posted online Oct. 4, 2019. We are grateful to Dr. Zachary L. Nimchuk for sharing the Nimchuk lab CRISPR system. We also thank Dr. Changfu Yao and Meyerowitz lab members for suggestions and discussions. The authors’ work was funded by the Howard Hughes Medical Institute and NASA grant NNX17AD53G to E.M.M. Author Contributions: T.L., A.Y., and E.M.M. conceived the experiments. T.L., A.Y. performed experiments. A.Y., T.L., and E.M.M. wrote the manuscript. Competing interests: A patent application (preliminary) has been filed based on this research.
Funders:
Funding AgencyGrant Number
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
NASANNX17AD53G
Record Number:CaltechAUTHORS:20191007-135703984
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20191007-135703984
Official Citation:Live imaging-assisted domain-specific CRISPR genome editing at single cell resolution in plants. Ting Li, An Yan, Elliot M. Meyerowitz. bioRxiv 793240; doi: https://doi.org/10.1101/793240
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:99118
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:07 Oct 2019 22:27
Last Modified:09 Mar 2020 13:19

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