State Key Laboratory of Protein and Plant Gene Research, School of Advanced Agricultural Sciences, Peking University, Beijing 100871, China.
Peking University Institute of Advanced Agricultural Sciences, Shandong Laboratory of Advanced Agricultural Sciences at Weifang, Shandong 261000, China.
Plant Commun. 2024 May 13;5(5):100823. doi: 10.1016/j.xplc.2024.100823. Epub 2024 Jan 18.
The inducible CRISPR activation (CRISPR-a) system offers unparalleled precision and versatility for regulating endogenous genes, making it highly sought after in plant research. In this study, we developed a chemically inducible CRISPR-a tool for plants called ER-Tag by combining the LexA-VP16-ER inducible system with the SunTag CRISPR-a system. We systematically compared different induction strategies and achieved high efficiency in target gene activation. We demonstrated that guide RNAs can be multiplexed and pooled for large-scale screening of effective morphogenic genes and gene pairs involved in plant regeneration. Further experiments showed that induced activation of these morphogenic genes can accelerate regeneration and improve regeneration efficiency in both eudicot and monocot plants, including alfalfa, woodland strawberry, and sheepgrass. Our study expands the CRISPR toolset in plants and provides a powerful new strategy for studying gene function when constitutive expression is not feasible or ideal.
诱导型 CRISPR 激活（CRISPR-a）系统为调控内源性基因提供了无与伦比的精确性和多功能性，因此在植物研究中备受关注。在这项研究中，我们通过将 LexA-VP16-ER 诱导系统与 SunTag CRISPR-a 系统相结合，开发了一种名为 ER-Tag 的植物化学诱导型 CRISPR-a 工具。我们系统地比较了不同的诱导策略，并实现了靶基因激活的高效率。我们证明了向导 RNA 可以进行多路复用和池化，用于大规模筛选有效的形态发生基因和参与植物再生的基因对。进一步的实验表明，这些形态发生基因的诱导激活可以加速再生，并提高双子叶植物和单子叶植物（包括紫花苜蓿、林地草莓和羊草）的再生效率。我们的研究扩展了植物中的 CRISPR 工具集，并提供了一种强大的新策略，用于研究基因功能，当组成型表达不可行或不理想时。
