Qin Xuejing, Hu Jing, Xu Guohui, Song Huifang, Zhang Lingyun, Cao Yibo
State Key Laboratory of Efficient Production of Forest Resources, Key Laboratory of Forest Silviculture and Conservation of the Ministry of Education, The College of Forestry, Beijing Forestry University, Beijing 100083, China.
College of Life and Health, Dalian University, Dalian 116000, China.
Plants (Basel). 2023 Aug 9;12(16):2905. doi: 10.3390/plants12162905.
The tumefaciens-mediated transformation for blueberries remains less efficient than is desirable. A new leaf callus regeneration and genetic transformation system was investigated in blueberries in this study. The leaf explants of 'Legacy' and 'Northland' were used to establish the stable callus induction system when placed on the woody plant medium (WPM) supplemented with 1.0 mg·L 2, 4-D, 0.4 mg·L 6-BA for 30 d; then, the callus was sub-cultured in the proliferation medium supplemented with 1.5 mg·L 2, 4-D, 0.4 mg·L 6-BA in the darkness at 25 °C every 30 days. The co-cultivation of callus with was operated on WPM plus 100 μM acetosyringone for 4 days; then, the transferred callus was grown in WPM supplemented with 1.5 mg·L 2,4-D, 0.4 mg·L 6-BA, 50 mg·L hygromycin, and 200 mg·L cefotaxime. The transgenic blueberry callus with both GFP signal and Hyg resistance was obtained from the transformed callus of 'Northland'. The rate of GFP signal detected in the transformed callus was as high as 49.02%, which was consistent with the PCR assay. Collectively, this study provides a highly efficient genetic transformation system in blueberry callus and a powerful approach for the molecular breeding of blueberries.
根癌农杆菌介导的蓝莓转化效率仍低于预期。本研究对蓝莓新的叶片愈伤组织再生和遗传转化系统进行了研究。以‘遗产’和‘北国’的叶片外植体为材料，置于添加1.0 mg·L 2,4 - D、0.4 mg·L 6 - BA的木本植物培养基（WPM）上30天，建立稳定的愈伤组织诱导体系；然后，将愈伤组织在添加1.5 mg·L 2,4 - D、0.4 mg·L 6 - BA的增殖培养基中于25℃黑暗条件下每30天继代培养一次。将愈伤组织与农杆菌在添加100 μM乙酰丁香酮的WPM上共培养4天；然后，将转接的愈伤组织在添加1.5 mg·L 2,4 - D、0.4 mg·L 6 - BA、50 mg·L潮霉素和200 mg·L头孢噻肟的WPM中培养。从‘北国’的转化愈伤组织中获得了具有绿色荧光蛋白（GFP）信号和潮霉素抗性的转基因蓝莓愈伤组织。在转化愈伤组织中检测到的GFP信号率高达49.02%，这与聚合酶链反应（PCR）检测结果一致。总体而言，本研究为蓝莓愈伤组织提供了一种高效的遗传转化系统，为蓝莓分子育种提供了有力方法。
