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Efficient Genome Editing in Rice Protoplasts Using CRISPR/CAS9 Construct

Abstract : Genome editing technologies, mainly CRISPR/CAS9, are revolutionizing plant biology and breeding. Since the demonstration of its effectiveness in eukaryotic cells, a very large number of derived technologies has emerged. Demonstrating and comparing the effectiveness of all these new technologies in entire plants is a long, tedious, and labor-intensive process that generally involves the production of transgenic plants and their analysis. Protoplasts, plant cells free of their walls, offer a simple, high-throughput system to test the efficiency of these editing technologies in a few weeks' time span. We have developed a routine protocol using protoplasts to test editing technologies in rice. Our protocol allows to test more than 30 constructs in protoplasts prepared from leaf tissues of 100, 9-11-day-old seedlings. CRISPR/CAS9 construct effectiveness can be clearly established within less than a week. We provide here a full protocol, from designing sgRNA to mutation analysis.
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https://hal.inrae.fr/hal-03328796
Contributor : Suzette Astruc <>
Submitted on : Monday, August 30, 2021 - 1:06:58 PM
Last modification on : Tuesday, August 31, 2021 - 3:27:51 AM

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Martine Bes, Leo Herbert, Thibault Mounier, Anne-Cécile Meunier, Franz Durandet, et al.. Efficient Genome Editing in Rice Protoplasts Using CRISPR/CAS9 Construct. Methods in Molecular Biology, 2238, pp.173-191, 2021, RICE GENOME ENGINEERING AND GENE EDITING, ⟨10.1007/978-1-0716-1068-8_11⟩. ⟨hal-03328796⟩

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