A simple and efficient CRISPR/Cas9 system permits ultra-multiplex genome editing in plants
A simple and efficient CRISPR/Cas9 system permits ultra-multiplex genome editing in plants
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一种简单有效的CRISPR/Cas9系统允许在植物中进行超多重基因组编辑
DOI：
10.1016/j.cj.2024.01.010
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发表时间：
2024-04-01
期刊：
影响因子：
6.6
通讯作者：
Cui，Xue'an
中科院分区：
文献类型：
--
作者：
Wu，Suting;Kyaw，Htin;Cui，Xue'an
CRISPR/Cas基因组编辑系统的发展和成熟为植物功能基因组学和遗传改良提供了有价值的工具。目前可用的基因组编辑工具的靶点数量有限，限制了它们在遗传研究中的应用。在这项研究中，我们开发了一种新的CRISPR/Cas9植物超多重基因组编辑系统，包括两个模板载体，八个供体载体，四个目的载体和一个引物设计软件包。通过组合Golden Gate克隆组装多个重复片段和Gateway重组组装大片段的优点，并通过改变用于组装sgRNA表达盒的扩增子的结构，植物超多重基因组编辑系统可以组装靶向超过40个基因组基因座的单个二元载体。组装了含有49个sgRNA表达盒的水稻敲除载体，并观察到高的共编辑效率。这种植物超多重基因组编辑系统推进了合成生物学和植物基因工程。
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.