粉红溲疏组培再生优化及遗传转化体系构建

doi:10.11707/j.1001-7488.LYKX20260041

林业科学 ›› 2026, Vol. 62 ›› Issue (9): 152-164.doi: 10.11707/j.1001-7488.LYKX20260041

• 研究论文 • 上一篇

粉红溲疏组培再生优化及遗传转化体系构建

郝英男¹,袁禹婷¹,屠雨欣¹,冯馨怡¹,张余周²,王慧慧²,吴普侠³,付春祥⁴,李进宇⁵,张鑫^1,*()

1. 陕西省林业综合重点实验室　旱区农业陕西实验室　西北农林科技大学林学院　杨凌 712100
2. 西北农林科技大学生命科学学院　杨凌 712100
3. 陕西省林业科学院　西安 710082
4. 中国科学院青岛生物能源与过程研究所　青岛 266101
5. 北京市园林绿化科学研究院　北京 100102

收稿日期:2026-01-20 修回日期:2026-05-16 出版日期:2026-09-10 发布日期:2026-09-16
通讯作者: 张鑫 E-mail:xin.zhang@nwafu.edu.cn
基金资助:
黄土高原植被恢复与土壤水文过程的互馈机理及调控策略项目（U24A20629）；国家重点研发计划项目（2024YFF306503）。

Optimization of Tissue Culture Regeneration and Construction of Genetic Transformation System of Deutzia rubens

Yingnan Hao¹,Yuting Yuan¹,Yuxin Tu¹,Xinyi Feng¹,Yuzhou Zhang²,Huihui Wang²,Puxia Wu³,Chunxiang Fu⁴,Jinyu Li⁵,Xin Zhang^1,*()

1. Key Comprehensive Laboratory of Forestry of Shaanxi Province　Shaanxi Laboratory of Dryland Agriculture　College of Forestry, Northwest A&F University　Yangling 712100
2. College of Life Sciences, Northwest A&F University　Yangling 712100
3. Shaanxi Academy of Forestry　Xi'an 710082
4. Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences　Qingdao 266101
5. Beijing Academy of Forestry and Landscape Architecture　Beijing 100102

Received:2026-01-20 Revised:2026-05-16 Online:2026-09-10 Published:2026-09-16
Contact: Xin Zhang E-mail:xin.zhang@nwafu.edu.cn

摘要/Abstract

摘要：

目的: 以实验室前期构建的2种溲疏属植物组培再生体系为基础，对溲疏属组培再生体系进行筛选和优化，并以最优再生体系为受体系统构建遗传转化体系，为重要园林绿化树种溲疏属植物的分子育种提供技术支撑，也为木本植物的从头驯化提供参考范式。方法: 以6种野生溲疏属植物的组培生根植株为研究对象，以形态学从上到下第1~5节叶片、叶柄、节间作为外植体，进行组培再生过程中基因型、外植体类型和成熟度的筛选，优化溲疏属植物组培再生体系。以最优再生体系为受体系统构建农杆菌介导的叶盘法遗传转化体系，并在该过程中对特美汀质量浓度进行筛选。结果: 受试的6种野生溲疏属植物中，再生效果最好的基因型为粉红溲疏；粉红溲疏组培再生的最适外植体为培养30天生根植株的形态学从上到下第1节展开叶片，分化率为100%；再生植株在1/2 MS +0.5 mg·L^–1 IBA生根培养基上的生根率为100%且植株和根系状态最好；生根植株在珍珠岩∶营养土=3∶7的炼苗移栽基质上状态最好，炼苗培养30天后，株高最高为15.68 cm；根癌农杆菌成功侵染粉红溲疏叶片并产生再生植株，在添加50 mg·L^–1特美汀的分化培养基中分化率最高达83.33%，遗传转化效率为16.7%。常规PCR检测确定外源基因Ruby以及NtGRF4-NtGIF1整合至粉红溲疏再生植株基因组中，且转基因植株呈现肉眼可见的红色表型。结论: 本研究建立以粉红溲疏为代表的溲疏属植物高效稳定组培再生体系，并首次构建农杆菌介导的叶盘法遗传转化体系，填补了溲疏属遗传转化技术空白。

关键词: 粉红溲疏, 组培再生, 叶盘法, 遗传转化, 从头驯化

Abstract:

Objective: On the basis of two previously established tissue culture regeneration systems of Deutzia species, the regeneration systems of Deutzia were screened and optimized, and a genetic transformation system was constructed with the optimal regeneration system as the recipient system in this study, aiming to provide technical support for molecular breeding of important landscape tree species in Deutzia, and a reference paradigm for the de novo domestication of woody plants. Method: The tissue culture rooted plantlets of six wild species in Deutzia were used as research materials. The leaves, petioles and internodes from the 1st to 5th node along the morphological axis were taken as explants. In the tissue culture regeneration process, the genotype, explant type and maturity were screened to optimize the tissue regeneration system of Deutzia. Subsequently, the optimized system used as the recipient system to construct an Agrobacterium tumefaciens-mediated leaf-disc genetic transformation system, and simultaneously screen the optimal concentration of Timentin. Result: Among the six tested wild Deutzia species, the genotype with the best regeneration was Deutzia rubens. The optimum explant for the tissue culture regeneration of D. rubens was the first expanded leaf of 30-day-old rooted plantlets, with a differentiation rate of 100%. The rooting rate of regenerated plantlets reached 100% on 1/2 MS medium supplemented with 0.5 mg·L^–1 IBA, showing the best growth of plantlets and root systems. The rooted plantlets grew best in the acclimatization and transplantation substrate with perlite∶nutrient soil = 3∶7. The maximum plant height was 15.68 cm after 30 days of acclimatization. A. tumefaciens successfully infected the leaves of D. rubens and regenerated plantlets. The differentiation rate reached the maximum of 83.33% at 50 mg·L^–1 Timentin, and the genetic transformation efficiency was 16.7%. Conventional PCR detection verified that exogenous Ruby and NtGRF4-NtGIF1 genes were integrated into the genome of regenerated D. rubens plants, and the transgenic plants showed a visible red phenotype. Conclusion: This study has established an efficient and stable tissue culture regeneration system of Deutzia represented by D. rubens, and an Agrobacterium-mediated leaf-disc genetic transformation system is for the first time constructed. The result fills the technical gap in genetic transformation of Deutzia, and provides core technical support for gene function analysis and targeted molecular improvement of this genus, as well as a practical reference for the utilization and de novo domestication breeding of wild woody plant resources.

Key words: Deutzia rubens, tissue culture regeneration, leaf-disc method, genetic transformation, de novo domestication

中图分类号:

S722.3⁺7

郝英男,袁禹婷,屠雨欣,冯馨怡,张余周,王慧慧,吴普侠,付春祥,李进宇,张鑫. 粉红溲疏组培再生优化及遗传转化体系构建[J]. 林业科学, 2026, 62(9): 152-164.

Yingnan Hao,Yuting Yuan,Yuxin Tu,Xinyi Feng,Yuzhou Zhang,Huihui Wang,Puxia Wu,Chunxiang Fu,Jinyu Li,Xin Zhang. Optimization of Tissue Culture Regeneration and Construction of Genetic Transformation System of Deutzia rubens[J]. Scientia Silvae Sinicae, 2026, 62(9): 152-164.

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引物名称 Primer name	引物序列 Primer sequences（5′→3′）	引物用途 Primer purpose
Ruby-F	CCACTGCGGATGGAACAGCATC	基因克隆与验证 Gene cloning and verification
Ruby-R	CTTGACGAACGTTGTCGAAACCG
NtGRF4-nF1	ATGAGTGCGACTACGATGAGGA
NtGRF4-nR1	CTTCGTTCCAGTGCTTCCAT

处理 Treatment	基因型 Genotype	外植体类型 Explant type	外植体成熟度 Explant maturity/node	分化率 Differentiation rate（%）	出芽数 Number of sprouts
I	粉红溲疏 D.rubens	叶片Leaf	1	100.00±0.00a	9.63±1.30b
			2	35.00±0.49cd	2.25±2.43f
			3~5	0.00±0.00f	0.00±0.00f
		叶柄Petiole	1	75.00±0.44b	5.38±2.45d
			2	25.00±0.44de	1.38±1.92fg
			3	10.00±0.31ef	0.38±1.92gh
			4	10.00±0.31ef	0.25±0.46gh
			5	0.00±0.00f	0.00±0.00h
		节间Internode	1~5	0.00±0.00f	0.00±0.00h
II	钩齿溲疏 D.baroniana	叶片Leaf	1	75.00±0.44b	16.5±0.73a
			2	10.00±0.32ef	4.15±0.88e
			3~5	0.00±0.00f	0.00±0.00h
		叶柄Petiole	1	5.23ef±0.23c	6.05±0.94cd
			2	5.88ef±0.32b	5.90±0.97cd
			3~5	0.00±0.00f	0.00±0.00h
		节间Internode	1	4.76ef±0.23c	6.75±0.64c
		节间Internode	2~5	0.00±0.00f	0.00±0.00h
III	褐毛溲疏 D.pilosa	叶片Leaf	1	75.00±0.45b	3.64±0.92e
		叶片Leaf	2	50.00±0.52c	1.18±0.40fgf
		叶柄Petiole	1~5	0.00±0.00f	0.00±0.00h
		节间Internode	1~5	0.00±0.00f	0.00±0.00h
IV	长江溲疏 D.schneideriana	叶片Leaf 叶柄Petiole 节间Internode	1~5	0.00±0.00f	0.00±0.00h
V	白溲疏 Deutzia albida	叶片Leaf 叶柄Petiole 节间Internode	1~5	0.00±0.00f	0.00±0.00h
VI	光萼溲疏 D.glabrata	叶片Leaf 叶柄Petiole 节间Internode	1~5	0.00±0.00f	0.00±0.00h

处理 Treatment	吲哚丁酸IBA/ （mg·L^–1）	生根率 Rootability （%）	根长 Root length/cm	株高 Plant height/cm
I	0.25	100a	4.23±0.50b	5.75±0.86b
II	0.5	100a	5.38±0.79a	7.09±0.93a
III	1	100a	5.19±0.92ab	5.93±0.78b
IV	2	100a	3.27±0.76c	4.50±0.85c

处理 Treatment	基质配比 Substrate ratio	成活率 Survival rate （%）	株高 Plant height/cm	植株状态 Healthy states of plant
Ⅰ	珍珠岩∶营养土=3∶7 Perlite∶nutrient soil =3∶7	100.00±0.00a	15.68±0.96a	叶片绿、长势好 Leaves remained green with robust growth
Ⅱ	珍珠岩∶营养土=6∶4 Perlite∶nutrient soil = 6∶4	93.75±0.25a	13.10±0.84b	叶片黄、长势较好 Leaves turned yellow and grew fairly well
Ⅲ	珍珠岩∶营养土=9∶1 Perlite∶nutrient soil = 9∶1	81.25±0.40a	5.92±0.53c	叶片黄、长势差 Leaves were yellow and growth was poor

处理 Treatment	特美汀 Timentin/ （mg·L^–1）	污染率 Contamination rate （%）	分化率 Differentiation rate （%）
I	50	0.00±0.00a	83.33±0.38a
II	100	0.00±0.00a	72.22±0.46a
III	200	0.00±0.00a	68.75±0.47a

粉红溲疏组培再生优化及遗传转化体系构建

Optimization of Tissue Culture Regeneration and Construction of Genetic Transformation System of Deutzia rubens

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