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- Title
A simple and efficient CRISPR/Cas9 system permits ultra-multiplex genome editing in plants.
- Authors
Suting Wua; Htin Kyaw; Zhijun Tong; Yirong Yang; Zhiwei Wang; Liying Zhang; Lihua Deng; Zhiguo Zhang; Bingguang Xiao; Quick, William Paul; Tiegang Lu; Guoying Xiao; Guannan Qin; Xue'an Cui
- Abstract
The development and maturation of the CRISPR/Cas genome editing system provides a valuable tool for plant functional genomics and genetic improvement. Currently available genome-editing tools have a limited number of targets, restricting their application in genetic research. In this study, we developed a novel CRISPR/Cas9 plant ultra-multiplex genome editing system consisting of two template vectors, eight donor vectors, four destination vectors, and one primer-design software package. By combining the advantages of Golden Gate cloning to assemble multiple repetitive fragments and Gateway recombination to assemble large fragments and by changing the structure of the amplicons used to assemble sgRNA expression cassettes, the plant ultra-multiplex genome editing system can assemble a single binary vector targeting more than 40 genomic loci. A rice knockout vector containing 49 sgRNA expression cassettes was assembled and a high co-editing efficiency was observed. This plant ultra-multiplex genome editing system advances synthetic biology and plant genetic engineering.
- Subjects
CRISPRS; GENOME editing; GENE expression; GENETIC engineering; BIOETHICS
- Publication
Crop Journal (2095-5421), 2024, Vol 12, Issue 2, p569
- ISSN
2095-5421
- Publication type
Article
- DOI
10.1016/j.cj.2024.01.010