Yang Yipeng, Chan Yuan, Wang Yongge, Guo Hao, Song Lina, Zhang Huali, Sun Liping, Cong Richen, Zhang Hua
Beijing Key Laboratory of Greening Plants Breeding, Beijing Academy of Forestry and Landscape Architecture, Beijing, China.
Front Plant Sci. 2024 Sep 5;15:1419313. doi: 10.3389/fpls.2024.1419313. eCollection 2024.
Bunge, belonging to the Acer genus in the Aceraceae family, is a commonly planted afforestation species across China, Japan, Korea, Europe, and North America. Renowned for its vibrant fall colors, it holds significant ecological and ornamental value.
In this study, ' Lihong ' was used as the research object. Starting from the callus induction of explants, the embryogenic callus of 'Lihong' was obtained by systematically optimizing the medium and culture conditions. Then, the candidate gene AtrGST894 screened by transcriptome sequencing was transformed into embryogenic callus by Agrobacterium-mediated transformation. The genetic transformation system of 'Lihong' embryogenic callus was initially established by continuously adjusting the conditions of Agrobacterium tumefaciens infection receptor materials, thus laying a material foundation for the study of the molecular regulation mechanism of 'Lihong' leaf color, and also preparing for the later molecular improvement breeding of . Therefore, this study has important theoretical and practical significance.
The results showed that the best medium for callus induction of was 1/2MS+2 mg/L 2,4-D+0.3 mg/L 6-BA+0.5 mg/L NAA; The embryogenic callus induction medium of was 1/2MS+3.0mg/L 6-BA+2.0mg/L TDZ+0.5mg/L IBA+0.1mg/L GA; The proliferation medium of embryogenic callus of was WPM+1.0mg/L TDZ+0.5mg/L IBA+0.1mg/L GA+3mg/L 6-BA+1.0mg/L KT; The infection experiment of Agrobacterium tumefaciens on the embryogenic callus of showed that the best antibacterial medium was WPM+30g/L sucrose+8g/L agar+0.5g/L acid-hydrolyzed casein+0.2mg/L KT+1.0 mg/L TDZ+0.5 mg/L IBA+0.1 mg/L GA+200mmol/L carboxybenzyl+200mg/L cephalosporin, and then WPM+30g/L sucrose+8g/L agar+0.5g/L acid-hydrolyzed casein+0.2mg/L KT+1.0 mg/L TDZ+0.5 mg/L IBA+0.1 mg/L GA+300mmol/L carboxybenzyl+200mg/L cephalosporin+25mg/L hygromycin. Screening medium screening, The obtained embryogenic callus browning rate, pollution rate and mortality rate were the lowest, and maintained vigorous growth.
The embryogenic callus was used as the infection material to verify that we successfully transferred the target gene into the embryogenic callus, which means that the genetic transformation system of embryogenic callus was partially completed, and the infection process could be effectively inhibited. Although there was partial browning, it could continue to proliferate. Therefore, in future experiments, the focus is still to continue to verify the optimal conditions for optimizing the genetic transformation of embryogenic callus and to solve the problems of difficulty in embryonic callus germination.
元宝枫属于槭树科槭属，是中国、日本、韩国、欧洲和北美洲广泛种植的造林树种。因其秋季色彩鲜艳而闻名，具有重要的生态和观赏价值。
本研究以‘丽红’元宝枫为研究对象。从外植体愈伤组织诱导开始，通过系统优化培养基和培养条件，获得‘丽红’元宝枫胚性愈伤组织。然后，将转录组测序筛选出的候选基因AtrGST894通过农杆菌介导转化法导入胚性愈伤组织。通过不断调整根癌农杆菌感染受体材料的条件，初步建立了‘丽红’元宝枫胚性愈伤组织的遗传转化体系，为‘丽红’元宝枫叶色分子调控机制的研究奠定了物质基础，也为后期的分子改良育种做准备。因此，本研究具有重要的理论和实践意义。
结果表明，‘丽红’元宝枫愈伤组织诱导的最佳培养基为1/2MS+2 mg/L 2,4-D+0.3 mg/L 6-BA+0.5 mg/L NAA；胚性愈伤组织诱导培养基为1/2MS+3.0mg/L 6-BA+2.0mg/L TDZ+0.5mg/L IBA+0.1mg/L GA；胚性愈伤组织增殖培养基为WPM+1.0mg/L TDZ+0.5mg/L IBA+0.1mg/L GA+3mg/L 6-BA+1.0mg/L KT；根癌农杆菌对‘丽红’元宝枫胚性愈伤组织的感染实验表明，最佳抑菌培养基为WPM+30g/L蔗糖+8g/L琼脂+0.5g/L酸水解酪蛋白+0.2mg/L KT+1.0 mg/L TDZ+0.5 mg/L IBA+0.1 mg/L GA+200mmol/L羧苄青霉素+200mg/L头孢霉素，然后是WPM+30g/L蔗糖+8g/L琼脂+0.5g/L酸水解酪蛋白+0.2mg/L KT+1.0 mg/L TDZ+0.5 mg/L IBA+0.1 mg/L GA+300mmol/L羧苄青霉素+200mg/L头孢霉素+25mg/L潮霉素。筛选培养基筛选，获得的胚性愈伤组织褐化率、污染率和死亡率最低，并保持旺盛生长。
以胚性愈伤组织作为感染材料，验证了我们成功地将目标基因导入胚性愈伤组织，这意味着‘丽红’元宝枫胚性愈伤组织的遗传转化体系部分完成，且感染过程能得到有效抑制。虽然有部分褐化，但仍能继续增殖。因此，在未来的实验中，重点仍将是继续验证优化‘丽红’元宝枫胚性愈伤组织遗传转化的最佳条件，并解决胚性愈伤组织萌发困难的问题。
