Efficient Regeneration Method and Pathway of in Vitro Leaves of Rhododendron

doi:10.11924/j.issn.1000-6850.casb2021-0104

Abstract

Abstract:

In vitro seedling leaves of six Rhododendron cultivars (‘Percy Wiseman’, ‘Cunningham's White’, ‘Halfdem Lem’, ‘Madame Masson’, ‘Scintillitation’ and ‘XXL’) were took as materials to establish efficient plant regeneration system. The effects of genotypes and exogenous hormones on the differentiation of in vitro leaves of six Rhododendron cultivars were studied, and the regeneration pathway was studied by morphology and histocytology. The results showed that TDZ was better than ZT in inducing the regeneration of in vitro leaves for the six Rhododendron cultivars. The differentiation rate of in vitro leaves induced by TDZ 0.04-0.08 mg/L was 100% and the average number of adventitious buds was 5.67-13 per leaf. The six Rhododendron cultivars could be used as candidate materials for genetic transformation, among which ‘Scintillition’, ‘Halfdem Lem’ and ‘Cunnigham’s White’ had stronger regeneration ability. Direct organ regeneration was the main regeneration pathway for the six Rhododendron cultivars, besides, there was also a few of leaves with callus indirect regeneration pathway to differentiate adventitious buds in ‘Percy Wiseman’ and ‘Madame Masson’ under TDZ induction. The results of the study could provide technical support and an experimental basis for the research on the mechanism of in vitro leaves differentiation of Rhododendron, and improve its plant regeneration frequency and genetic transformation efficiency.

Key words: Rhododendron, in vitro leaves, efficient regeneration, histocytology, regeneration pathway

CLC Number:

S603.8

Peng Lvchun, Liao Duosi, Jie Weijia, Song Jie, Cai Yanfei, Zhang Lu, Li Shifeng. Efficient Regeneration Method and Pathway of in Vitro Leaves of Rhododendron[J]. Chinese Agricultural Science Bulletin, 2021, 37(34): 77-83.

Figures/Tables 6

References 25

[1]	解玮佳, 王继华, 卢红春, 等. 21个高山杜鹃引进品种的亲本分析[J]. 河南农业大学学报, 2015, 49(5):622-627.
[2]	李志斌, 白霄霞, 李萍, 等. 国际合作架金桥世界名花放异彩——高山杜鹃引进开发研究十年成就回顾[J]. 农业科技通讯, 2010(13):12-15.
[3]	兰熙, 张乐华, 张金政, 等. 杜鹃花属植物育种研究进展[J]. 园艺学报, 2012, 39(9):1829-1838.
[4]	刘闵豪, 徐郡儡, 叶靖, 等. 农杆菌介导的杜仲叶片愈伤组织遗传转化体系[J]. 林业科学, 2020, 56(2):79-88.
[5]	马春华. 枣叶片再生途径的组织学研究及影响遗传转化因素的分析[D]. 郑州:河南农业大学, 2011: 6.
[6]	向发云, 韩永超, 曾祥国, 等. ‘晶瑶’草莓高效再生及遗传转化体系的建立[J]. 中国农学通报, 2017, 33(15):36-42.
[7]	白宇清, 蓝伟根, 谢利娟. 杜鹃组织培养研究进展[J]. 南方林业科学, 2016, 44(4):25-28.
[8]	吴福建, 李凤兰, 黄凤兰, 等. 杜鹃花研究进展[J]. 东北农业大学学报, 2008, 39(1):139-144.
[9]	张开文, 从睿, 王定跃. 国内杜鹃属植物组培快繁技术的研究进展[J]. 林业科技, 2020, 45(1):29-33.
[10]	杜凤国, 李伟伟, 王欢, 等. 兴安杜鹃基本培养基和外植体选择的初步研究[J]. 安徽农业科学院, 2019, 47(12):136-138.
[11]	陈春福. 福建杜鹃花属若干种类的离体繁殖与保存及其ISSR分析[D]. 福州:福建农林大学, 2012:15-18.
[12]	郭颖. 三种高山杜鹃组织培养快繁技术研究[D]. 北京:北京林业大学, 2015:11-13.
[13]	Zaytseva Y G, Poluboyarova T V, Novikova T I. Effects of thidiazuron on in vitro morphogenic response of Rhododendron sichotense Pojark. and Rhododendron catawbiense cv. Grandiflorum leaf explants[J]. In Vitro Cellular & Developmental Biology- Plant, 2016, 52(1):56-63.
[14]	刘燕, 孙超, 王济红, 等. 桃叶杜鹃愈伤组织诱导与增殖研究[J]. 种子, 2014, 33(3):35.
[15]	汪玲敏, 李世峰, 解玮佳, 等. 云南杜鹃(Rhododendron yunnanense Franch.)无菌叶片高频再生体系的建立[J]. 分子植物育种, 2015, 13(8):1877-1883.
[16]	吕秀立, 魏翔莺, 尹丽娟, 等. 大叶常绿杜鹃叶片离体培养及不定芽再生[J]. 南京林业大学学报:自然科学版, 2014, 38(2):165-168.
[17]	Pavingerova D. The influence of thidiazuron on shoot regeneration from leaf explants of fifteen cultivars of Rhododendron[J]. Biologia Plantarum, 2009, 53(4):797-799. doi: 10.1007/s10535-009-0147-3 URL
[18]	皮明雪, 张少敏, 郭涛, 等. 甘蓝型油菜小孢子培养中基因型及处理条件对成苗的影响[J]. 种子, 2020, 39(9):13-17.
[19]	吴晓军, 胡喜贵, 陈向东, 等. 小麦成熟胚再生体系优化效果评价及高再生率基因型筛选[J]. 江苏农业科学, 2019, 47(11):74-77.
[20]	Guo B, Abbasi B H, Zeb A, et al. Thidiazuron: A multi-dimensional plant growth regulator[J]. African Journal of Biotechnology, 2011, 10(45):8984-9000. doi: 10.5897/AJB URL
[21]	刘淼, 曹后男, 宗成文, 等. TDZ对牛皮杜鹃叶片分化及继代增殖的影响[J]. 西北农业学报, 2012, 21(12):158-162.
[22]	苏家乐, 刘晓青, 何丽斯, 等. 杜鹃品种‘江南春早’叶片离体再生体系的建立[J]. 分子植物育种2019, 17(4):1283-1289.
[23]	罗琳, 白洁, 陈超, 等. 美容杜鹃叶片再生及愈伤组织耐热生理研究[J]. 西北植物学报, 2014, 34(7):1377-1382.
[24]	Vejsadova H, Pretova A. Somatic embryogenesis in Rhododendron catawbiense ‘Grandiflorum’[J]. Acta Hot, 2003, 6(16):467-470.
[25]	罗彭, 庄平, 白洁. 大白杜鹃、美容杜鹃和喇叭杜鹃的组织培育[J]. 植物生理学通讯, 2007, 43(2):326.

处理编号	TDZ	ZT	NAA
1	0	0	0.01
2	0.04	0	0.01
3	0.06	0	0.01
4	0.08	0	0.01
5	0.1	0	0.01
6	0.15	0	0.01
7	0.2	0	0.01
8	0	1	0.01
9	0	2	0.01
10	0	3	0.01
11	0	4	0.01

处理编号	TDZ	ZT	NAA
1	0	0	0.01
2	0.04	0	0.01
3	0.06	0	0.01
4	0.08	0	0.01
5	0.1	0	0.01
6	0.15	0	0.01
7	0.2	0	0.01
8	0	1	0.01
9	0	2	0.01
10	0	3	0.01
11	0	4	0.01

处理编号	D4	W10	W17	W22	W29	X
1	14.29±0d	14.29±0e	14.29±0e	14.29±0f	19.05±4.76c	19.05±4.76e
2	76.19±4.76b	100.00±0a	100.00±0a	80.95±4.76b	100.00±0a	100.00±0a
3	85.71±0b	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
4	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
5	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
6	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
7	100.00±0a	100.00±0a	100.00±0a	95.24±4.76a	100.00±0a	100.00±0a
8	14.29±8.25d	28.57±0d	33.33±4.76d	42.86±0e	71.43±0b	38.10±4.76d
9	42.86±8.25c	38.10±4.76c	47.62±4.76c	52.38±4.76d	76.19±4.76b	76.19±4.76c
10	42.86±8.25c	85.71±8.25b	71.43±0b	66.67±4.76c	100.00±0a	90.47±4.76b
11	47.62±4.76c	100.00±0a	95.24±4.76a	71.43±0c	100.00±0a	100.00±0a

处理编号	D4	W10	W17	W22	W29	X
1	14.29±0d	14.29±0e	14.29±0e	14.29±0f	19.05±4.76c	19.05±4.76e
2	76.19±4.76b	100.00±0a	100.00±0a	80.95±4.76b	100.00±0a	100.00±0a
3	85.71±0b	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
4	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
5	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
6	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a	100.00±0a
7	100.00±0a	100.00±0a	100.00±0a	95.24±4.76a	100.00±0a	100.00±0a
8	14.29±8.25d	28.57±0d	33.33±4.76d	42.86±0e	71.43±0b	38.10±4.76d
9	42.86±8.25c	38.10±4.76c	47.62±4.76c	52.38±4.76d	76.19±4.76b	76.19±4.76c
10	42.86±8.25c	85.71±8.25b	71.43±0b	66.67±4.76c	100.00±0a	90.47±4.76b
11	47.62±4.76c	100.00±0a	95.24±4.76a	71.43±0c	100.00±0a	100.00±0a

处理	D4	W10	W17	W22	W29	X
1	0.00±0f	0.00±0f	0.00±0e	0.00±0e	0.00±0h	0.00±0e
2	4.33±0.333c	11.67±0.882ab	7.33±0.667b	7.33±0.333a	6.33±0.333c	4.33±0.333bc
3	6.33±0.333a	13.00±0.577a	9.67±0.333a	7.67±0.333a	9.33±0.a	5.67±0.333a
4	5.33±0.333b	10.67±0.667bc	7.00±0.577b	5.67±0.333b	7.67±0.333b	5.33±0.333ab
5	5.33±0.333b	11.67±0.667ab	8.00±0.577b	5.00±0.577bc	6.33±0.333c	4.67±0.333abc
6	5.67±0.333a	9.67±0.333c	5.33±0.333c	5.67±0.333b	6.67±0.333c	4.00±0.577c
7	5.33±0.333b	4.67±0.333d	5.67±0.333c	4.67±0.333c	6.67±0.333c	4.33±0.333bc
8	0.00±0f	0.00±0f	0.00±0e	0.00±0e	2.33±0.333f	0.33±0.333e
9	0.33±0.333f	1.00±0.577f	1.67±0.333d	0.67±0.333e	4.67±0.333de	1.67±0.333d
10	1.33±0.333e	2.67±0.333e	2.33±0.333d	1.67±0.333d	5.33±0.333d	2.33±0.333d
11	3.33±0.333d	3.67±0.333de	2.33±0.333d	2.33±0.333d	4.33±0.333e	3.67±0.333c