利用 RNA 病毒载体传递 CRISPR-Cas 进行植物免转化基因组编辑的方案。

IF 1.3 Q4 BIOCHEMICAL RESEARCH METHODS

STAR Protocols Pub Date : 2024-12-20 Epub Date: 2024-11-05 DOI:10.1016/j.xpro.2024.103437

Huanhuan Lou, Haiying Xiang, Wanli Zeng, Jiarui Jiang, Jianduo Zhang, Li Xu, Chenglu Zhao, Qian Gao, Zhenghe Li

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引用次数: 0

摘要

植物病毒载体已成为 CRISPR-Cas 试剂递送的理想工具。在这里，我们介绍了一种利用工程化 RNA 病毒载体瞬时递送 CRISPR-Cas 成分进行无 DNA 植物基因组编辑的方案。我们介绍了病毒载体构建、通过农用接种恢复病毒载体、机械接种目标植物宿主、体细胞诱变频率分析和突变植株再生等步骤。该方法编辑效率高，无需进行稳定的植物转化。有关该方案使用和执行的完整细节，请参阅 Liu 等人的文章1。

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Protocol for transformation-free genome editing in plants using RNA virus vectors for CRISPR-Cas delivery.

Plant virus vectors have emerged as promising tools for CRISPR-Cas reagent delivery. Here, we present a protocol for DNA-free plant genome editing using an engineered RNA virus vector for the transient delivery of CRISPR-Cas components. We describe steps for viral vector construction, viral vector recovery through agroinoculation of Nicotiana benthamiana, mechanical inoculation of target plant hosts, analysis of somatic mutagenesis frequency, and regeneration of mutant plants. The method achieves high editing efficiency and eliminates the need for stable plant transformation. For complete details on the use and execution of this protocol, please refer to Liu et al.¹.

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