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Abstract:Agrobacterium-mediated transformation is an important approach for genetic modification of wheat, but it still suffers from several unresolved critical technical bottlenecks, including low transformation efficiency, strong genotype dependence, severe browning of calli, and low regeneration efficiency. In this study, immature embryos of the model cultivar Fielder were used as explants to systematically investigate the effects of embryo age, acetosyringone (AS) concentration and Agrobacterium infection duration, phosphinothricin (PPT) selection pressure, and exogenous hormone (abscisic acid, ABA; kinetin, KT) ratios on callus induction, regeneration and transformation efficiency. Using the optimized protocol, applicability was evaluated in the cultivars Yangmai 20, Zhengmai 7698, and Kenong 199. The results showed that embryos harvested 12 days after anthesis exhibited the lowest browning rate (4.40%) and were most suitable as recipients for transformation. When the Agrobacterium suspension was adjusted to OD600=0.5, supplementation with 200 μmol·L-1 AS and a 10 min infection produced the highest induction rate of resistant green callus spots (34.29%). Addition of 0.5 mg·L-1 ABA to the induction medium and 1.5 mg·L-1 KT to the regeneration medium significantly enhanced embryogenic callus formation and plant regeneration. The optimal PPT selection concentration was determined to be 10 mg·L-1. Under the optimized conditions, transformation efficiency of Fielder increased significantly from 8.16% before optimization to 14.83%; transformation efficiencies of Yangmai 20, Zhengmai 7698, and Kenong 199 reached 9.03%, 7.32% and 8.70%, respectively, approaching or exceeding the pre-optimization level of Fielder. In summary, the optimized system established in this study improves wheat genetic transformation efficiency and is broadly applicable across different genetic backgrounds, providing a technical platform for gene functional analysis and molecular breeding in wheat.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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