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CRISPR-Cas9-mediated construction of a cotton CDPK mutant library for identification of insect-resistance genes

Plant Communications 2024年第11期5卷 23-40页

作者： Fuqiu Wang Sijia Liang Guanying Wang Tianyu Hu Chunyang Fu Qiongqiong Wang Zhongping Xu Yibo Fan lianlian che Ling Min Bo Li Lu Long Wei Gao Xianlong Zhang Shuangxia Jin Hubei Hongshan Laboratory National Key Laboratory of Crop Genetic ImprovementHuazhong Agricultural UniversityWuhan 430070China Academy of Industry Innovation and Development Huanghuai UniversityZhumadianHenan 463000China Xinjiang Key Laboratory of Crop Biotechnology Institute of Nuclear and Biological TechnologyXinjiang Academy of Agricultural SciencesUrumqi 830091 XinjiangChina National Key Laboratory of Cotton Bio-breeding and Integrated Utilization School of Life ScienceHenan UniversityHenan 475004China These authors contributed equally to this article

Calcium-dependent protein kinases(CDPKs)act as key signal transduction enzymes in plants,especially in response to diverse stresses,including *** this study,a comprehensive analysis of the CDPK gene family in upland cotton revealed that GhCPKs are widely expressed in multiple cotton tissues and respond positively to various biotic and abiotic *** developed a strategy for screening insect-resistance genes from a CRISPR-Cas9 mutant library of *** library was created using 246 single-guide RNAs targeting the GhCPK gene family to generate 518 independent T0 *** average target-gene coverage was 86.18%,the genome editing rate was 89.49%,and the editing heritability was 82%.An insect bioassay in the field led to identification of 14 GhCPK mutants that are resistant or susceptible to *** mutant that showed the clearest insect resistance,cpk33/74(in which the homologous genes GhCPK33 and GhCPK74 were knocked out),was selected for further *** secretions from Spodoptera litura induced a rapid influx of Ca^(2+) in cpk33/74 leaves,resulting in a significant increase in jasmonic acid content.S-adenosylmethionine synthase is an important protein involved in plant stress response,and protein interaction experiments provided evidence for interactions of GhCPK33 and GhCPK74 with GhSAMS1 and *** addition,virus-induced gene silencing of GhSAMS1 and GhSAM2 in cotton impaired defense against *** study demonstrates an effective strategy for constructing a mutant library of a gene family in a polyploid plant species and offers valuable insights into the role of CDPKs in the interaction between plants and herbivorous insects.

关键词： cotton CDPKs mutant library CRISPR-Cas9 Ca^(2+)influx insect resistance

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