莲的离体快速繁殖技术

doi:10.11983/CBB21020

植物学报 ›› 2021, Vol. 56 ›› Issue (5): 605-613.DOI: 10.11983/CBB21020

莲的离体快速繁殖技术

熊雅倩¹^,², 邓显豹¹^,³, 张会会¹, 杨东¹^,³, 孙恒¹, 刘娟¹, 杨美¹^,³^,^*()

¹中国科学院武汉植物园植物种质创新与特色农业重点实验室, 武汉 430074
²中国科学院大学, 北京 100049
³中国科学院核心植物园(武汉), 协同中心, 武汉 430074

收稿日期:2021-01-26 接受日期:2021-05-27 出版日期:2021-09-01 发布日期:2021-08-31
通讯作者: 杨美
作者简介:* E-mail: yangmei815815@wbgcas.cn
基金资助:
中国科学院前沿科学重点研究计划(QYZDB-SSW-SMC017);中国科学院青年创新促进会项目(2017390);国家自然科学基金(31772353);国家自然科学基金(31872136)

In Vitro Rapid Propagation of Nelumbo nucifera

Yaqian Xiong¹^,², Xianbao Deng¹^,³, Huihui Zhang¹, Dong Yang¹^,³, Heng Sun¹, Juan Liu¹, Mei Yang¹^,³^,^*()

¹Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan 430074, China
²University of Chinese Academy of Sciences, Beijing 100049, China
³Center of Economic Botany, Core Botanical Gardens, Chinese Academy of Sciences, Wuhan 430074, China

Received:2021-01-26 Accepted:2021-05-27 Online:2021-09-01 Published:2021-08-31
Contact: Mei Yang

摘要/Abstract

摘要： 以莲(Nelumbo nucifera)授粉后18天的莲子胚芽为外植体, 通过初代培养、继代培养和炼苗移栽, 建立了莲离体快速繁殖体系。结果表明, 将胚芽外植体诱导出无菌苗的最适初代培养基为MS固体培养基添加0.5 mg∙L^-1 6-BA、0.5 mg∙L^-1 NAA、30 g∙L^-1蔗糖、0.5 g∙L^-1活性炭和0.8 g∙L^-1琼脂, 培养60天诱导率高于85%, 其中秋红阳走茎节数最多(3.9)。最佳继代培养基为将初代培养基中的蔗糖浓度提高到80 g∙L^-1, 走茎采用两节一切的分苗切法, 无菌苗可50天继代1次, 最多可继代6次, 不同品种的增殖系数介于4.0-6.7之间, 以秋红阳最高(6.7)。于5-7月将生根的走茎无菌苗移栽入泥炭:塘泥=1:2 (v/v)的混合基质中进行培养, 成活率均大于83.9%。采用上述快繁技术, 理论上1个莲子胚芽经过近1年可繁殖出种苗1 465株。该研究建立了莲的离体快繁技术体系, 可为莲种苗的规模化生产提供技术支持。

关键词: 莲, 无菌苗, 快速繁殖, 植株再生

Abstract: Using fresh embryos isolated from developing lotus seeds of 18 days after pollination as explants, a rapid propagation system of lotus was established through primary culture, subculture plantlets hardening and transplantation. Results showed that the primary culture medium of MS+0.5 mg·L^-1 6-BA+0.5 mg·L^-1 NAA+30 g·L^-1 sucrose+0.5 g·L^-1 activated carbon+0.8 g·L^-1agar was the optimum for the explant growth. The explant induction rate was up to 85% after cultured for 60 days, Qiuhongyang had the highest amount of stem nodes. In the subculture, the optimal concentration of sucrose in the medium was 80 g·L^-1. When dividing aseptic seedling for subculture, cutting with two stem nodes showed the highest multiplication coefficient. The multiplication coefficient of different varieties was ranged from 4.0 to 6.7, and Qiuhongyang was the highest (6.7). Aseptic seedlings were suggested to be subcultured every 50 days, with high multiplication rates for up to 6 rounds of subcultures. The rooted plantlets were transplanted to pots containing medium of peat:pond soil=1:2 (v/v) during May to July, and the survival rate of transplanted plants was higher than 83.9%. Based on these results, using this in vitro propagation system, it is estimated that 1 465 seedlings be developed from a single lotus seed within one year. This study has established a rapid in vitro propagation system for lotus, which provides the basis for the factory scale production of lotus plantlets.

Key words: lotus, aseptic seedlings, rapid propagation, plant regeneration

熊雅倩, 邓显豹, 张会会, 杨东, 孙恒, 刘娟, 杨美. 莲的离体快速繁殖技术. 植物学报, 2021, 56(5): 605-613.

Yaqian Xiong, Xianbao Deng, Huihui Zhang, Dong Yang, Heng Sun, Juan Liu, Mei Yang. In Vitro Rapid Propagation of Nelumbo nucifera. Chinese Bulletin of Botany, 2021, 56(5): 605-613.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB21020

https://www.chinbullbotany.com/CN/Y2021/V56/I5/605

图/表 8

表1 初代培养基组成成分

Table 1 The component of the primary culture medium

Types	NAA (mg∙L^-1)	6-BA (mg∙L^-1)	Sucrose (g∙L^-1)	Activated charcoal (g∙L^-1)
I	0.5	0.5	30	0.5
II	0.5	1	30	0.5
III	0.5	0.5	80	0.5
IV	0.5	0.5	30	1

表2 继代培养基的激素配比

Table 2 The combinations of plant hormones in subculture medium

Types	NAA (mg∙L^-1)	6-BA (mg∙L^-1)
1	0.5	0.5
2	0.3	0.5
3	0.1	0.5
4	0.5	1.5
5	1	0.5
6	0.3	1

图1 秋红阳授粉后18天莲子胚芽的初代培养 (A) 授粉后18天的莲子胚芽外植体; (B) 接种后培养1天的胚芽; (C) 初代培养10天的无菌苗长根; (D) 初代培养60天的健壮走茎苗。Bars=1 cm

Figure 1 Primary culture of Qiuhongyang immature embryo explants from developing seeds of 18 days after pollination (A) Immature embryo explants from developing seeds of 18 days after pollination; (B) Immature embryo cultured one day after inoculation; (C) The rooted plantlets at 10 days after primary culture; (D) The aseptic seedling at 60 days after primary culture. Bars=1 cm

表3 培养基组分对秋红阳无菌苗初代培养的影响(接种60天)

Table 3 Effects of medium components on the growth of Qiuhongyang aseptic seedlings in the primary culture (60 days after inoculation)

Types	The amount of inoculation	Rooting rate (%)	The amount of nodes	The amount of apical bud	The amount of leaves
I	331	100	3.9±0.36 a	0.9±0.03 a	4.3±0.48 a
II	134	98	4.0±0.05 a	0.7±0.04 b	3.8±0.27 a
III	122	95	3.2±0.19 b	0.7±0.01 b	4.3±0.13 a
IV	300	95	3.6±0.33 ab	0.9±0.13 a	3.9±0.22 a

图2 不同莲品种对无菌苗初代培养的走茎节数(A)和诱导率(B)的影响不同小写字母表示差异显著(P<0.05)。

Figure 2 Effects of lotus varieties on the number of stem nodes (A) and the induction rate (B) of lotus aseptic seedlings in the primary culture Different lowercase letters indicate significant differences (P<0.05).

图3 无菌苗继代培养繁殖系数和死亡率 (A) 不同走茎切割方式对秋红阳无菌苗继代培养繁殖系数和死亡率的影响(x: 2节, y: 1节加1个生长点, z: 1节, w: 1个生长点); (B) 不同激素配比对秋红阳无菌苗继代培养繁殖系数和死亡率的影响, 激素配比(编号1-6)见表2; (C) 不同品种对无菌苗继代培养繁殖系数和死亡率的影响。不同小写字母表示差异显著(P<0.05)。

Figure 3 The multiplication coefficient and mortality rate of subculture of aseptic seedlings (A) Effect of dividing methods in Qiuhongyang aseptic seedlings on the multiplication coefficient and mortality rate of subculture (x: Cuttings with two nodes, y: Cuttings with one node and one growing tip, z: Cuttings with one nodes, w: Cuttings with one growing tip); (B) Effects of plant hormone combinations in Qiuhongyang on the multiplication coefficient and mortality rate of subculture, combinations of plant hormones (No.1-6) are in Table 2; (C) Effect of lotus varieties on the multiplication coefficient and mortality rate of subculture. Different lowercase letters indicate significant differences at P<0.05.

图4 秋红阳无菌苗炼苗和移栽 (A1), (A2) 无菌苗继代培养50天; (B1), (B2) 移栽10天; (C1) 移栽40天长出立叶; (C2) 移栽40天的地下茎; (C3) 正常繁殖所得地下茎。(A1), (A2), (B1), (B2) Bars=1 cm, (C1)-(C3) Bars=10 cm

Figure 4 Hardening and transplanting of Qiuhongyang aseptic seedlings (A1), (A2) Aseptic seedlings at 50 days of subculture; (B1), (B2) Plantlets at 10 days after transplanting; (C1) Standing leaves started to emerge at 40 days after transplanting; (C2) Rhizomes at 40 days after transplanting; (C3) Rhizome taken from plants grew in ponds. (A1), (A2), (B1), (B2) Bars=1 cm, (C1)-(C3) Bars=10 cm

表4 不同品种无菌苗移栽存活率及叶片数

Table 4 Effects of lotus varieties on the survival rate and the number of leaves developed after transplanting

Lotus varieties	Survival rate (%)		The amount of expanded leaves		The amount of standing leaves
Lotus varieties	10 d	20 d	10 d	20 d	50 d
Qiuhongyang	93.7±3.0 a	83.9±1.6 a	0.4±0.10 b	1.8±0.07 b	0.7±0.04 a
Qiurihonghua	93.3±6.7 a	93.3±6.7 a	0.7±0.07 a	2.7±0.15 a	0.5±0.04 ab
WBG_S1	95.8±7.2 a	93.3±6.3 a	0.5±0.00 b	1.7±0.05 b	0.3±0.06 b
Baihuajianlian	87.8±6.3 a	87.2±18.0 a	0.5±0.11 b	2.5±0.40 a	0.5±0.13 ab

参考文献 32

[1]	蔡颖欣, 汤宇环, 邵晓宇, 黄霞 (2017). 莲子胚培养再生植株及原生质体分离的研究. 种子 36(3), 125-127, 134.
[2]	何碧珠, 曾明星, 赵时端, 王家福, 赖钟雄 (2002). 建莲茎尖离体培养研究初报. 福建农林大学学报(自然科学版) 31, 59-61.
[3]	黄宁珍, 付传明, 赵志国, 唐凤鸾, 石云平 (2010). 桂林小花苣苔离体快速繁殖技术. 植物学报 45, 744-750. DOI
[4]	柯卫东, 彭静, 刘玉平, 黄新芳 (2001). 试管藕诱导技术研究. 武汉植物学研究 19, 173-175.
[5]	孔德政, 李艳妮, 杨秋生, 刘广甫 (2007). 荷花胚组织培养的初步研究. 河南科学 25, 593-595.
[6]	李峰, 周雄祥, 柯卫东, 黄新芳, 朱红莲, 钟兰, 宗义湘, 吴曼, 彭静, 李双梅, 袁田垚 (2020). 湖北省莲产业发展调研报告. 湖北农业科学 59(23), 101-106, 109.
[7]	刘建平, 王芳, 杜彩娴, 梁少丽, 曾莉莎, 郑芝波 (2016). 荷花幼胚组织培养技术研究. 现代农业科技 (15), 140, 142.
[8]	刘义满, 柯卫东 (2012). 关于提高莲产业效益的建议. 长江蔬菜 (16), 134-137.
[9]	彭燕, 张玲莉, 杨小青, 阳静, 李娜, 宋金春 (2017). 莲子心总生物碱对人肝癌细胞的抑制作用. 中国药师 20, 1009-1012.
[10]	唐凤鸾, 赵健, 赵志国, 夏科, 仇硕 (2019). 走马胎的组织培养与快速繁殖. 植物学报 54, 378-384. DOI
[11]	唐嘉瓅, 邱杰, 黄学辉 (2020). 基因组学技术大发展助力园艺植物研究取得新进展. 植物学报 55, 1-4. DOI
[12]	王其超, 张行言 (1998). 二元分类法在荷花品种分类中的应用. 北京林业大学学报 20(2), 33-37.
[13]	王其超, 张行言 (2005). 中国荷花品种图志. 北京: 中国林业出版社. pp. 34-43.
[14]	徐君, 李静会, 李欣, 周玉珍, 韦庆华, 姜红卫 (2013). 荷花顶芽初代组织培养. 江苏农业科学 41(3), 38-39.
[15]	岳建华, 董艳, 王小画, 孙佩霞, 王思颖, 张新年, 张琰 (2020). 早花百子莲叶片器官发生和胚胎发生再生体系的建立. 植物学报 55, 588-595.
[16]	曾明星, 何碧珠, 罗银华 (2005). 建莲藕茎尖离体培养快繁技术及应用. 福建农业科技 (3), 13-14.
[17]	张建福, 王锋 (2002). 莲藕组织培养与微繁殖技术初探. 上海农业科技 (6), 17-18.
[18]	张文婷, 何燕红, 舒宁, 邢景景, 刘宝骏, 包满珠, 刘国锋 (2019). 金黄花滇百合植株再生与离体快繁技术体系的建立. 植物学报 54, 773-778. DOI
[19]	赵芹, 李效尊, 徐国鑫, 阴筱, 尹静静, 吴修 (2016). 莲组织培养与分子生物学研究进展. 分子植物育种 14, 1587-1594.
[20]	Abe T, Futsuhara Y (1986). Genotypic variability for callus formation and plant regeneration in rice ( Oryza sativa L.). Theor Appl Genet 72, 3-10. DOI PMID
[21]	Arunyanart S, Chaitrayagun M (2005). Induction of somatic embryogenesis in lotus ( Nelumbo nucifera Geartn.). Sci Horticult 105, 411-420. DOI URL
[22]	Buathong R, Saetiew K, Phansiri S, Parinthawong N, Arunyanart S (2013). Tissue culture and transformation of the antisense DFR gene into lotus (Nelumbo nucifera Gaertn.) through particle bombardment. Sci Horticult 161, 216-222. DOI URL
[23]	Deng XB, Xiong YQ, Li J, Yang D, Liu J, Sun H, Song HY, Wang YM, Ma JY, Liu YL, Yang M (2020). The establishment of an efficient callus induction system for lotus ( Nelumbo nucifera). Plants 9, 1436. DOI URL
[24]	Guo HB (2009). Cultivation of lotus ( Nelumbo nucifera Gaertn. ssp. nucifera) and its utilization in China. Genet Resour Crop Evol 56, 323-330. DOI URL
[25]	Jun MY, Karki R, Paudel KR, Sharma BR, Adhikari D, Kim DW (2016). Alkaloid rich fraction from Nelumbo nucifera targets VSMC proliferation and migration to suppress restenosis in balloon-injured rat carotid artery. Atherosclerosis 248, 179-189. DOI URL
[26]	La-ongsri W, Trisonthi C, Balslev H (2009). Management and use of Nelumbo nucifera Gaertn. in Thai wetlands. Wetlands Ecol Manage 17, 279-289. DOI URL
[27]	Liu QQ, Zhang DS, Liu FL, Qin M, Tian DK (2019). Micropropagation of Nelumbo nucifera ‘Weishan Hong’ through germfree mature embryos. In Vitro Cell Dev Biol Plant 55, 305-312. DOI URL
[28]	Mahmad N, Taha RM, Othman R, Saleh A, Hasbullah NA, Elias H (2014). Effects of NAA and BAP, double-layered media, and light distance on in vitro regeneration of Nelumbo nucifera Gaertn. (lotus), an aquatic edible plant. The Scientific World J 2014, 745148.
[29]	Shou SY, Miao LX, Zai WS, Huang XZ, Guo DP (2008). Factors influencing shoot multiplication of lotus ( Nelumbo nucifera). Biol Plantarum 52, 529-532. DOI URL
[30]	Sridhar KR, Bhat R (2007). Lotus-A potential nutraceutical source. J Agric Technol 3, 143-155.
[31]	Yang M, Zhu LP, Pan C, Xu LM, Liu YL, Ke WD, Yang PF (2015). Transcriptomic analysis of the regulation of rhizome formation in temperate and tropical lotus ( Nelumbo nucifera). Sci Rep 5, 13059. DOI PMID
[32]	Zhang XY, Wang XY, Wu TT, Li BX, Liu TQ, Wang R, Liu Q, Liu ZJ, Gong YQ, Shao CS (2015). Isoliensinine induces apoptosis in triple-negative human breast cancer cells through ROS generation and p38 MAPK/JNK activation. Sci Rep 5, 12579. DOI PMID

莲的离体快速繁殖技术

In Vitro Rapid Propagation of Nelumbo nucifera

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