Development of Fragrant Thermosensitive Genic Male Sterile Line Rice Using CRISPR/Cas9

作者：Tengkui Chen, Na Pu, Min Ni, Huabin Xie, Zhe Zhao, Juan Hu, Zhanhua Lu, Wuming Xiao, Zhiqiang Chen, Xiuying He, Hui Wang · 发表于：Agronomy · 年份：2025 · DOI：10.3390/agronomy15020411 · 被引用次数：1 · 研究领域：GABA and Rice Research、Microbial Metabolic Engineering and Bioproduction、Medicinal Plants and Neuroprotection

This study aimed to develop an aromatic thermosensitive genic male sterile (TGMS) line in indica rice using CRISPR/Cas9 technology. The TMS5 and FGR in the high-quality conventional rice variety Huahang 48 were targeted for editing using CRISPR/Cas9 technology. CRISPR/Cas9 vectors designed for TMS5 and FGR were constructed and introduced into rice calli through Agrobacterium-mediated transformation. Transgenic seedlings were subsequently regenerated, and the target sites of the edited plants were analyzed via sequencing. A total of fifteen T0 double mutants were successfully obtained. Three mutants without T-DNA insertion were screened in the T1 generation by the PCR detection of hygromycin gene fragments, and homozygous mutants without T-DNA insertion were screened in the T2 generation by the sequencing analysis of the mutation sites, named Huahang 48s. Huahang 48s exhibited complete sterility at 24 °C and pollen transfer at 23 °C. The 2-acetyl-1-pyrroline (2-AP) content was detected in the young panicles, leaves, and stems of Huahang 48s. The leaves of Huahang 48s had the highest 2-AP content, contrasting with the absence of 2-AP in HuaHang 48. F1 hybrids that crossed Huahang 48s with two high-quality restorer lines were superior to the two parents in terms of yield per plant and 1000-grain weight. Huahang 48s has a certain combining ability and application potential in two-line cross breeding. The successful application of CRISPR/Cas9 technology in Huahang 48 established a...