Promoter editing represents an innovative approach to introduce quantitative trait variation (QTV) in crops. However, an efficient promoter editing system for QTV needs to be established. Here we develop a CRISPR–Cas12a promoter editing (CAPE) system that combines a promoter key-region estimating model and an efficient CRISPR–Cas12a-based multiplexed or singular editing system. CAPE is benchmarked in rice to produce QTV continuums for grain starch content and size by targeting OsGBSS1 and OsGS3, respectively. We then apply CAPE for promoter editing of OsD18, a gene encoding GA3ox in the gibberellin biosynthesis pathway. The resulting lines carry a QTV continuum of semidwarfism without significantly compromising grain measures. Field trials demonstrated that the OsD18 promoter editing lines have the same yield performance and antilodging phenotype as the Green Revolution OsSD1 mutants in different genetic backgrounds. Hence, promoter editing of OsD18 generates a quantitative Green Revolution trait. Together, we demonstrate a CAPE-based promoter editing and tuning pipeline for efficient production of useful QTV continuum in crops.
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Sequence conservation data were downloaded from PlantRegMap. The TF binding motif datasets were downloaded from JASPAR (2020 Core Plants collections). Genetic variations and phenotype data of rice were downloaded from MBKbase. The rice reference genome (MSU/Tigr7) was downloaded from the Rice Genome Annotation Project. The tomato reference genome (SL4.0) was downloaded from the Solanaceae Genomics Network. The maize reference genome (AGPv4) was downloaded from MaizeGDB. Public open chromatin and histone modification datasets were retrieved from the NCBI under the accession numbers GSE26610 and GSM2084219 for rice, GSE164297 and PRJNA381300 for tomato, and PRJNA599454 and PRJNA417726 for maize.
The instructions of feature data processing and the complete pipeline of CAPE for calculating aggregate scores and detecting KRs have been deposited to GitHub (https://github.com/zhangtaolab/CAPE).
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We thank Q. Qian (China National Rice Research Institute, Chinese Academy of Agricultural Sciences) for providing xiaowei, a OsD18-null mutant in a slightly different Nipponbare background. This research was supported by the National Natural Science Foundation of China (award nos. 32270433, 32101205, 32072045 and 31960423) to Y. Zhang, X.Z. and X.T.; and the National Agricultural Major Science and Technology Program of China (award no. NK2022010204) to Y. Zhang and J.Z.; the Technology Innovation and Application Development Program of Chongqing (award no. CSTC2021JSCX-CYLHX0001) to X.T., J.Z. and Y. Zhang; the Sichuan Science and Technology Program (award nos. 2021JDRC0032, 2021YFH0084 and 2021YFYZ0016) to J.Z. and Y. Zhang; and the Open Foundation of Jiangsu Key Laboratory of Crop Genetics and Physiology (award no. YCSL202009) to J.Z., Y. Zhang and T.Z. It was also supported by the NSF Plant Genome Research Program (grant nos. IOS-2029889 and IOS-2132693) and the MAES Competitive Grants Program (CGP) to Y.Q.
The authors declare no competing interests.
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Supplementary Figs. 1–23.
Supplementary Table 1. The differential expression of rice genes via mRNA-Seq. Table 2. Gibberellin-response genes in rice. Table 3. Gibberellin biosynthesis genes in rice. Table 4. Comparison of promoter editing efficiency in rice. Table 5. Oligonucleotides used in this study.
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Zhou, J., Liu, G., Zhao, Y. et al. An efficient CRISPR–Cas12a promoter editing system for crop improvement. Nat. Plants 9, 588–604 (2023). https://doi.org/10.1038/s41477-023-01384-2