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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Crop Improvement and Genetics Research » Research » Publications at this Location » Publication #418263
Research Project: Molecular Resources for Enhanced Crop Biotechnology

Location: Crop Improvement and Genetics Research

Title: Novel agrobacterium fabrum str. 1D1416 for citrus transformation

Author
item ALABED, DIAA - Oak Ridge Institute For Science And Education (ORISE)
item Tibebu, Redeat
item Ariyaratne, Menaka
item SHAO, MIN - Oak Ridge Institute For Science And Education (ORISE)
item Milner, Matthew
item Thomson, James

Submitted to: Microorganisms
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/30/2024
Publication Date: 9/30/2024
Citation: Alabed, D., Tibebu, R.A., Ariyaratne, M.D., Shao, M., Milner, M.J., Thomson, J.G. 2024. Novel agrobacterium fabrum str. 1D1416 for citrus transformation. Microorganisms. 12(10). https://doi.org/10.3390/microorganisms12101999.
DOI: https://doi.org/10.3390/microorganisms12101999

Interpretive Summary: In this study, we screen a collection of wild Agrobacterium strains for efficient citrus transformation and identify the novel strain 1D1416, which shows a high capacity for transforming various citrus species. We further modified this strain to include the GAANTRY system, enabling assembly and stable maintenance of large stacked gene constructs within the T-DNA. This novel strain, combined with a modified transformation protocol, offers an improved method for citrus species transformation.

Technical Abstract: Citrus is one of the world's most important and widely produced fruit crops, with 166.3 million tons harvested from over 10.6 million hectares in 2022. Challenges in crop maintenance, produc-tion, and fruit quality necessitate developing new traits through Agrobacterium-mediated genetic transformation. While a few Agrobacterium strains (EHA105, GV3101, LBA4404) are known to transform citrus, many wild strains remain untested. We screened 41 wild-type Agrobacterium strains isolated from various woody species and identified five capable of DNA transfer into citrus cells. Strain 1D1416 demonstrated the highest transient transformation frequency in Car-rizo epicotyl explants (88%), outperforming the control EHA105 (84%) with comparable shoot regeneration rates (32% and 42%, respectively). Notably, 1D1416 exhibited no overgrowth and had the lowest necrosis and mortality rates in transformed tissues. It efficiently transferred the DsRed gene and induced galls in mature tissues of Mexican lime (70%), lemon (48%), Washington navel orange (25%), and clementine (6%). Genome sequencing of 1D1416 allowed disarming of the native T-DNA and addition of GAANTRY technology. This novel strain, combined with an optimized transformation procedure, make it a valuable tool for advancing citrus transfor-mation.