伊藤杂种‘和谐’组培快繁体系的建立

doi:10.11983/CBB23105

植物学报 ›› 2024, Vol. 59 ›› Issue (3): 441-451.DOI: 10.11983/CBB23105

伊藤杂种‘和谐’组培快繁体系的建立

康敏¹^,⁴, 张美莹¹^,⁴, 齐秀双⁶, 佟宁宁⁴^,⁵, 李旸⁴^,⁵, 舒庆艳⁴^,⁵, 刘政安⁴^,⁵, 吕长平²^,³^,^*(), 彭丽平⁴^,⁵^,^*()

¹湖南农业大学园艺学院, 长沙 410128
²湖南农业大学风景园林与艺术设计学院, 长沙 410128
³湖南农业大学湖南省中亚热带优质花木繁育与利用工程技术研究中心, 长沙 410128
⁴中国科学院植物研究所植物多样性与特色经济作物全国重点实验室, 北京 100093
⁵国家植物园, 北京 100093
⁶天香源(辽宁)生物科技有限责任公司, 葫芦岛 125300

收稿日期:2023-08-03 接受日期:2023-12-19 出版日期:2024-05-01 发布日期:2023-12-29
通讯作者: *吕长平, 湖南农业大学副教授, 硕士生导师, 主要研究方向为园林与观赏园艺植物栽培生理。曾获湖南省科技进步三等奖4次, 国家教学成果二等奖、湖南农业大学教学成果二等奖各1次; 以通讯作者和第一作者身份在Horticultural Science and Technology及湖南农业大学学报(自然科学版)等学术期刊上发表研究论文30多篇。E-mail: changpinglv@sina.com;彭丽平, 中国科学院植物研究所副研究员。长期从事牡丹群体遗传、新品种培育、组织快繁以及有效成分功能评价等研究。以通讯作者和第一作者身份在Horticulture Research、Physiologia Plantarum和Foods等国际主流学术期刊上发表研究论文10余篇。pengliping@ibcas.ac.cn
基金资助:
国家自然科学基金青年科学基金(32201609);中国科学院植物研究所企业横向项目(E0119C6001)

Establishment of a Fast Breeding System for Itoh Hybrid ‘He Xie’ in Tissue Culture

Min Kang¹^,⁴, Meiying Zhang¹^,⁴, Xiushuang Qi⁶, Ningning Tong⁴^,⁵, Yang Li⁴^,⁵, Qingyan Shu⁴^,⁵, Zheng’an Liu⁴^,⁵, Changping Lü²^,³^,^*(), Liping Peng⁴^,⁵^,^*()

¹College of Horticulture, Hunan Agricultural University, Changsha 410128, China
²College of Landscape Architecture and Art Design, Hunan Agricultural University, Changsha 410128, China
³Hunan Provincial Engineering and Technology Research Center for Breeding and Utilization of Central-subtropical High-quality Flowering Trees, Hunan Agricultural University, Changsha 410128, China
⁴National Key Laboratory of Plant Diversity and Special Economic Crops, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China
⁵National Botanical Garden, Beijing 100093, China
⁶Tian Xiang Yuan (Liaoning) Biotechnology Co. Ltd., Huludao 125300, China

Received:2023-08-03 Accepted:2023-12-19 Online:2024-05-01 Published:2023-12-29
Contact: *E-mail: changpinglv@sina.com; pengliping@ibcas.ac.cn

摘要/Abstract

摘要： 以伊藤杂种‘和谐’(Itoh hybrids ‘He Xie’)的鳞芽为材料建立离体快繁技术体系, 可克服传统方法繁育较慢的缺点, 加速伊藤杂种优良品种的繁育推广。采用单因子试验设计, 分别探究不同灭菌时间、不同植物生长调节剂浓度、不同根诱导时间以及不同生根苗等级对‘和谐’组培苗启动、增殖、生根和驯化效果的影响。结果表明, 用2%次氯酸钠溶液的最佳灭菌时间为12分钟, 鳞芽污染率为9.09%; 最佳初始培养基配方为MS+1.5 mg∙L^-1 6-BA+0.2 mg∙L^-1 GA₃+0.5 mg∙L^-1 AgNO₃; 最佳增殖培养基配方为MS+450 mg∙L^-1 CaCl₂+0.5 mg∙L^-1 6-BA+0.2 mg∙L^-1 IBA+0.2 mg∙L^-1 GA₃+0.5 mg∙L^-1 AgNO₃, 增殖系数为3.3; 无根苗在根诱导培养基1/2MS+1.0 mg∙L^-1腐胺+2.0 mg∙L^-1 IBA上, 经4°C低温暗培养8天后常温光照培养30天, 再转入根形成培养基1/2MS+1.0 g∙L^-1 AC, 培养20天生根率达66.7%; 苗移栽的基质为珍珠岩:蛭石:草炭土=1:1:1 (v/v/v), 移栽60天后, 一级苗成活率最高为52.0%, 二级苗和三级苗则大部分死亡, 表明生根质量对移栽成活至关重要。

关键词: 伊藤‘和谐’, 启动与增殖, 生根, 鳞芽, 移栽驯化

Abstract: An in vitro rapid propagation system using scale buds of Itoh hybrids ‘He Xie’ can overcome the shortcomings of the slow traditional breeding methods and promote the adoption of the excellent Itoh hybrid varieties. In this study, we used the buds of ‘He Xie’ as explants to investigate the effects of various factors including disinfection time, plant growth regulators (PGRs) concentration and root induction time, and different rooted seedling grades on the initiation, proliferation, rooting and domestication of ‘He Xie’ with one-way experimental design. Our results showed: the optimal disinfection time of buds by 2% sodium hypochlorite solution was 12 min, with a contamination rate of 9.09%; the optimal initial culture medium was MS+1.5 mg∙L^-1 6-BA+0.2 mg∙L^-1 GA₃+0.5 mg∙L^-1AgNO₃; the optimal proliferation culture medium was MS+450 mg∙L^-1 CaCl₂+0.5 mg∙L^-1 6-BA+0.2 mg∙L^-1 IBA+0.2 mg∙L^-1GA₃+0.5 mg∙L^-1 AgNO₃ with a proliferation rate of 3.3. Rootless seedlings were cultured on root induction medium 1/2MS+1.0 mg∙L^-1putrescine+2.0 mg∙L^-1 IBA for 8 d at 4°C in dark and then 30 d at room temperature under light, and finally transferred to root formation medium 1/2MS+1.0 g∙L^-1 AC for 20 d, a rooting rate of 66.7% was observed. The rooted seedlings were transplanted on a growing matrix of perlite:vermiculite:charcoal soil=1:1:1 (v/v/v) for 60 d, with the highest transplant survival rate of 52.0% being observed for first-grade rooted seedlings, but most of the second- and third-grade seedlings being dead, suggesting that the quality of rooting is critical for transplant survival.

Key words: Itoh ‘He Xie’, initiation and proliferation, rooting, scale buds, transplant domestication

康敏, 张美莹, 齐秀双, 佟宁宁, 李旸, 舒庆艳, 刘政安, 吕长平, 彭丽平. 伊藤杂种‘和谐’组培快繁体系的建立. 植物学报, 2024, 59(3): 441-451.

Min Kang, Meiying Zhang, Xiushuang Qi, Ningning Tong, Yang Li, Qingyan Shu, Zheng’an Liu, Changping Lü, Liping Peng. Establishment of a Fast Breeding System for Itoh Hybrid ‘He Xie’ in Tissue Culture. Chinese Bulletin of Botany, 2024, 59(3): 441-451.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB23105

https://www.chinbullbotany.com/CN/Y2024/V59/I3/441

图/表 7

图1 ‘和谐’鳞芽启动与单芽增殖 (A) 花; (B) 外植体; (C)-(E) 鳞芽在不同浓度(0.5、1.0和1.5 mg∙L-1) 6-BA启动培养30天后试管苗生长状态; (F) 增殖培养30天后单芽生长状态。Bars=2 cm

Figure 1 Bud initiation and single bud proliferation of 'He Xie' (A) Flowers; (B) Explants; (C)-(E) The performance of test-tube plantlets after bud initiation at different 6-BA concentrations (0.5, 1.0, and 1.5 mg∙L-1) for 30 days; (F) Growth situation of single shoot after proliferation culture for 30 days. Bars=2 cm

图2 ‘和谐’试管苗生根和驯化移栽的生长表现 (A) 用于生根诱导的单芽(bar=2 cm); (B) 春化30天(冷处理8天, 根诱导30天, 根伸长20天后于4°C冷藏30天)的生根苗(bar= 2 cm); (C)-(F) 不同生根质量的试管苗(C: 三级苗; D: 二级苗; E, F: 一级苗) (bars=2 cm); (G)-(I) 驯化60天不同生根质量的移栽苗(G: 三级苗; H: 二级苗; I: 一级苗) (bars=3 cm)

Figure 2 The growth performance for rooting and domesticated transplanting of ‘He Xie’ tube plantlet (A) Single shoots used for root induction (bar=2 cm); (B) Rooted seedlings after 30 d of vernalization (cold treatment for 8 days, root induction for 30 days, and root elongation for 20 days, cold storage at 4°C for 30 days) (bar=2 cm); (C)-(F) The test tube plantlets with different rooting qualities (C: Tertiary; D: Secondary; E, F: Primary) (bars=2 cm); (G)-(I) The transplanted seedlings of different rooting quality for domesticating 60 d (G: Tertiary; H: Secondary; I: Primary) (bars=3 cm)

表1 不同消毒时间下‘和谐’鳞芽的污染率和存活率

Table 1 Contamination rate and surviveal rate of ‘He Xie’ buds treated with different disinfection times

Treatments	Time (min)	Sodium hypochlorite	Contamination rate (%)	Survival rate (%)
X1	8	2%	62.5	100
X2	10	2%	40.0	100
X3	12	2%	9.09	100

表2 不同浓度6-BA对‘和谐’鳞芽初代培养的影响

Table 2 Effect of different 6-BA concentrations on the primary culture of ‘He Xie’ buds

Treatments	GA₃(mg∙L^-1)	6-BA (mg∙L^-1)	Induction rate (%)	Browning rate (%)	Multiplication factor (n)	Stem length (cm)	Leaves per plant (n)
1	0.2	0.5	100	20	2.40±0.26 b	2.58±0.60 a	6.47±1.01 a
2	0.2	1.0	100	10	2.80±0.10 a	2.95±0.23 a	7.80±0.70 a
3	0.2	1.5	100	10	2.93±0.12 a	3.05±0.41 a	6.93±0.21 a

表3 GA3浓度对‘和谐’试管苗增殖的影响

Table 3 Effect of GA3 concentration on proliferation of ‘He Xie’ test-tube plantlets

Treatments	6-BA (mg∙L^-1)	IBA (mg∙L^-1)	GA₃(mg∙L^-1)	Multiplication factor (n)	Browning rate (%)
1	0.5	0.2	0.2	3.25±0.33 a	0.04±0.07 a
2	0.5	0.2	0.5	2.67±0.29 ab	0.08±0.07 a
3	0.5	0.2	0.8	2.13±0.33 b	0.17±0.07 a

表4 IBA浓度与诱导时间对诱导生根的影响

Table 4 Effect of IBA concentration and induction time on induced rooting

Treatments	IBA (mg∙L^-1)	Induction time (d)	Rooting rate (%)	Rooting number (n)	Root length (cm)
1	0.5	30	1.59±2.75 h	0.67±1.15 cd	1.53±2.66 bc
2	0.5	40	19.05±2.75 fg	2.00±1.15 abc	5.20±2.66 a
3	0.5	50	14.29±0.00 g	1.33±0.58 cde	3.87±0.23 ab
4	1.0	30	19.05±8.24 fg	1.33±0.58 cde	5.33±1.94 a
5	1.0	40	3.17±5.50 h	0.67±1.15 cd	1.53±2.66 bc
6	1.0	50	0.00±0.00 h	0.00±0.00 d	0.00±0.00 c
7	1.5	30	14.29±0.00 g	1.67±1.15 abc	4.17±2.75 ab
8	1.5	40	33.33±8.25 de	2.00±0.50 abc	4.73±0.13 a
9	1.5	50	28.57±0.00 ef	2.67±1.15 ab	5.33±0.50 a
10	2.0	30	66.67±8.25 a	3.33±0.58 a	4.77±0.65 a
11	2.0	40	42.86±0.00 bcd	2.78±0.39 ab	5.33±0.62 a
12	2.0	50	47.62±8.24 bc	2.33±0.67 abc	4.95±1.58 a
13	2.5	30	38.10±8.25 cde	1.72±0.25 abc	4.85±1.73 a
14	2.5	40	71.43±0.00 a	2.06±0.42 abc	5.26±0.52 a
15	2.5	50	52.38±8.24 b	2.11±0.84 abc	5.38±0.98 a

表5 不同生根数与生根质量对不同时间移栽苗成活率的影响

Table 5 Effect of different rooting number and rooting quality on the survival rate of transplanted seedlings at different time

Rooting quality	Survival rate (%)
Rooting quality	10 d	20 d	30 d	40 d	50 d	60 d
Level 1	96.00	92.00	72.00	72.00	64.00	52.00
Level 2	87.80	85.37	73.17	62.85	60.98	48.78
Level 3	52.38	47.62	28.57	19.05	14.29	14.29

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伊藤杂种‘和谐’组培快繁体系的建立

Establishment of a Fast Breeding System for Itoh Hybrid ‘He Xie’ in Tissue Culture

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