Rapeseed (Brassica napus) is the world’s third most important oilseed crop and a key beneficiary of heterosis, or hybrid vigor. Male sterility systems are essential to hybrid seed production, with cytoplasmic male sterility (CMS) and genic male sterility (GMS) being the most widely used. While CMS is cost-effective, it requires complex restoring–maintaining relationships, whereas GMS is stable but limited by difficulties in producing fully sterile populations. Existing solutions, such as seed production technology (SPT), depend on transgenic markers and face safety concerns from potential gene flow. Based on these challenges, there is an urgent need for a reliable, visually trackable, and efficient system to improve hybrid breeding.
A research team from Huazhong Agricultural University and Hubei Hongshan Laboratory has introduced a new strategy for hybrid seed production in rapeseed. Their study, published (DOI: 10.1093/hr/uhae270) on September 25, 2024, in Horticulture Research, details how targeted mutation of BnaMS1 and BnaMS2 genes, combined with the RUBY visual reporter, enables a simple two-line hybrid system. By creating male-sterile lines through CRISPR/Cas9 and coupling them with a transgenic maintainer identifiable by red pigmentation, the researchers demonstrated a reliable method for large-scale hybrid seed propagation without the laborious processes typical of CMS and traditional GMS approaches.
The study focused on overcoming barriers in recessive genic male sterility (RGMS) systems by developing sterile lines directly from elite rapeseed varieties. Using CRISPR/Cas9, the researchers successfully disrupted BnaMS1 and BnaMS2, producing plants that exhibited complete male sterility yet retained normal growth and female fertility. Unlike natural or chemically induced mutations, this gene-editing approach ensured precise, stable, and reproducible results across diverse parental backgrounds.
To address the challenge of distinguishing maintainer and sterile plants, the team tested three reporter systems: DsRed, BnaA07.PAP2, and RUBY. Among these, RUBY—known for generating stable red pigmentation—proved the most effective marker. The PMR (pigment-based) system provided a clear visual cue, enabling rapid screening without specialized equipment. Maintainer lines carrying RUBY exhibited a 1:1 segregation pattern when crossed with sterile lines, producing easily identifiable pink (fertile) and green (sterile) seedlings within days of germination. This simplified sorting process reduces cost, time, and reliance on sophisticated fluorescence technology.
Overall, the integration of CRISPR/Cas9 mutagenesis with RUBY-based maintainers represents a powerful advance for rapeseed breeding. It ensures efficient propagation of RGMS plants and provides a scalable framework adaptable to other Brassica crops, enhancing the accessibility of hybrid seed technology.
“Our findings present a practical solution to one of the long-standing challenges in rapeseed hybrid seed production,” said Dr. Chuchuan Fan, senior author of the study. “By combining CRISPR/Cas9 with a simple visual marker, we have established a two-line system that is stable, efficient, and easily scalable. The RUBY reporter enables rapid identification at the seedling stage, eliminating costly and complex sorting processes. This method not only accelerates hybrid breeding in rapeseed but also offers a promising model for other crops where male sterility systems are essential for hybrid seed production.”
This innovation has wide-ranging applications in crop breeding and food security. For rapeseed, the new two-line system simplifies hybrid seed production, lowers costs, and ensures stable propagation of male-sterile lines. The approach reduces dependence on cytoplasmic sterility and minimizes risks linked to transgenic pollen escape. Beyond rapeseed, the method is readily transferable to other Brassica crops such as cabbage, mustard, and broccoli, potentially accelerating hybrid breeding across the vegetable and oilseed industries. By offering breeders a reliable tool for large-scale seed production, this technology could enhance global agricultural productivity and resilience, contributing to sustainable food systems in the face of rising demand.
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References
DOI
10.1093/hr/uhae270
Original Source URL
https://doi.org/10.1093/hr/uhae270
Funding information
Funding for this research was obtained from various sources, including the Technology Major Project on Key Techniques of Agricultural Biological Breeding (2023ZD0404203), the Key Research Projects of Hubei Province (2022BBA0039), the National Natural Science Foundation of China (32172021, 31371240, 31671279), the Wuhan Science and Technology Major Project on Key Techniques of Biological Breeding and Breeding of New Varieties (grant no. 2022021302024851), and the Open Project of Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs, PR China (KF2023006).
About Horticulture Research
Horticulture Research is an open access journal of Nanjing Agricultural University and ranked number one in the Horticulture category of the Journal Citation Reports ™ from Clarivate, 2023. The journal is committed to publishing original research articles, reviews, perspectives, comments, correspondence articles and letters to the editor related to all major horticultural plants and disciplines, including biotechnology, breeding, cellular and molecular biology, evolution, genetics, inter-species interactions, physiology, and the origination and domestication of crops.