1. Construction of Host Plant Insect‐Resistance Mutant Library by High‐Throughput CRISPR/Cas9 System and Identification of A Broad‐Spectrum Insect Resistance Gene
- Author
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Lin Sun, Muna Alariqi, Yaxin Wang, Qiongqiong Wang, Zhongping Xu, Muhammad Naeem Zafar, Guangqin Yang, Ruoyu Jia, Amjad Hussain, Yilin Chen, Xiao Ding, Jiawei Zhou, Guanying Wang, Fuqiu Wang, Jianying Li, Jiawei Zou, Xiangqian Zhu, Lu Yu, Yiwen Sun, Sijia Liang, Fengjiao Hui, Luo Chen, Weifeng Guo, Yanqin Wang, Huaguo Zhu, Keith Lindsey, Xinhui Nie, Xianlong Zhang, and Shuangxia Jin
- Subjects
cotton ,CRISPR/Cas9 ,GhEPS15 ,GhMLP423 ,high‐throughput genome editing ,plant‐insect interaction ,Science - Abstract
Abstract Insects pose significant challenges in cotton‐producing regions. Here, they describe a high‐throughput CRISPR/Cas9‐mediated large‐scale mutagenesis library targeting endogenous insect‐resistance‐related genes in cotton. This library targeted 502 previously identified genes using 968 sgRNAs, generated ≈2000 T0 plants and achieved 97.29% genome editing with efficient heredity, reaching upto 84.78%. Several potential resistance‐related mutants (10% of 200 lines) their identified that may contribute to cotton‐insect molecular interaction. Among these, they selected 139 and 144 lines showing decreased resistance to pest infestation and targeting major latex‐like protein 423 (GhMLP423) for in‐depth study. Overexpression of GhMLP423 enhanced insect resistance by activating the plant systemic acquired resistance (SAR) of salicylic acid (SA) and pathogenesis‐related (PR) genes. This activation is induced by an elevation of cytosolic calcium [Ca2+]cyt flux eliciting reactive oxygen species (ROS), which their demoted in GhMLP423 knockout (CR) plants. Protein‐protein interaction assays revealed that GhMLP423 interacted with a human epidermal growth factor receptor substrate15 (EPS15) protein at the cell membrane. Together, they regulated the systemically propagating waves of Ca2+ and ROS, which in turn induced SAR. Collectively, this large‐scale mutagenesis library provides an efficient strategy for functional genomics research of polyploid plant species and serves as a solid platform for genetic engineering of insect resistance.
- Published
- 2024
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