A Modular Cloning Toolkit for Genome Editing in Cereals

B - Book chapters etc edited externally

Hahn, F., Sanjurjo-Loures, L. and Nekrasov, V. 2022. A Modular Cloning Toolkit for Genome Editing in Cereals. in: Bilichak, A. and Laurie, J. D. (ed.) Accelerated Breeding of Cereal Crops New York, NY Humana Press Inc. pp. 209-224

AuthorsHahn, F., Sanjurjo-Loures, L. and Nekrasov, V.
EditorsBilichak, A. and Laurie, J. D.

The CRISPR/Cas technology has recently become the tool of choice for gene editing applications in plants.Here we describe the method of assembling CRISPR/Cas DNA constructs, using the modular cloning (MoClo), for deployment in cereal species. For this purpose, we use a cloning toolkit that consists of a library of Golden Gate (GG) modules, which encode genetic elements, such as promoters, coding sequences of CRISPR/Cas nucleases (e.g. Cas9), terminators, guide RNA backbones, etc. A part of the toolkit are modulesthat allow assembly of tRNA-sgRNA polycistronic units without a PCR step involved. The tRNA-sgRNA system has proved to be efficient in a number of plant species, including cereals (rice, wheat). The cloning toolkit we present can be used for a variety of gene editing applications, including gene deletions and single nucleotide substitutions. It can also be easily expanded by adding new standardised parts.

KeywordsCRISPR; Cas9; Plant; Genome editing; Golden Gate; MoClo; Cereal; Wheat
Page range209-224
Year of Publication2022
Book titleAccelerated Breeding of Cereal Crops
PublisherHumana Press Inc
Place of publicationNew York, NY
SeriesSpringer Protocols Handbooks
Digital Object Identifier (DOI)https://doi.org/10.1007/978-1-0716-1526-3_10
Web address (URL)https://link.springer.com/protocol/10.1007/978-1-0716-1526-3_10
FunderBiotechnology and Biological Sciences Research Council
Funder project or codeDesigning Future Wheat (DFW) [ISPG]
Open accessPublished as non-open access
Output statusPublished
Publication dates
Online21 Oct 2021

Permalink - https://repository.rothamsted.ac.uk/item/986z9/a-modular-cloning-toolkit-for-genome-editing-in-cereals

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