‘京枣39’离体多倍体诱导及其增殖、生根培养体系优化

赵汗青; 王强; 庞晓明; 薄文浩; 李颖岳

doi:10.12171/j.1000-1522.20230039

‘京枣39’离体多倍体诱导及其增殖、生根培养体系优化

1.
林木遗传育种全国重点实验室，林木花卉遗传育种教育部重点实验室，北京林业大学生物科学与技术学院，北京林业大学，北京 100083
2.
内蒙古自治区林业和草原种苗总站，内蒙古呼和浩特 010051

基金项目: 国家自然科学基金项目（32271925），中央高校基本科研业务费专项（QNTD202305）。

详细信息

作者简介:
赵汗青。主要研究方向：经济林良种繁育。Email：hanqingzhao@bjfu.edu.cn　地址：100083 北京市海淀区清华东路35号北京林业大学生物科学与技术学院

责任作者:
李颖岳，教授，博士生导师。主要研究方向：经济林良种繁育。Email：yingyueli@bjfu.edu.cn　地址：同上。

中图分类号: S665.1
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出版历程
- 收稿日期: 2023-02-23
- 修回日期: 2023-03-13
- 录用日期: 2024-05-09
- 网络出版日期: 2024-05-13
- 刊出日期: 2024-06-29

In vitro polyploid induction and optimization of its proliferation and rooting culture system for ‘Jingzao 39’

1.
State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Genetics and Breeding in Forest Trees and Omamental Plants, Beijing Laboratory of Urban and Rural Ecological Environment, School of Biological Sciences and Biotechnology, Beiing Forestry University, Beijing 100083, China
2.
Inner Mongolia General Station of Seed and Seedling of Forestry and Grassland, Hohhot 010051, Inner Mongolia, China

摘要

摘要:
目的
研究秋水仙素溶液质量浓度、处理时间和预培养时间对‘京枣39’离体多倍体诱导的影响，优化枣树良种‘京枣39’的增殖和生根培养基，建立‘京枣39’的离体多倍体诱导体系，获得多倍体植株。
方法
采用完全随机区组试验设计，探索不同秋水仙素溶液质量浓度和处理时间组合对‘京枣39’不定芽再生的影响，不同预培养时间对‘京枣39’离体多倍体诱导的影响，不同生长调节剂组合对‘京枣39’增殖培养的影响，不同基本培养基和生长调节剂组合对‘京枣39’组培苗生根培养的影响。
结果
（1）当秋水仙素溶液质量浓度为80 mg/L、处理48 h时‘京枣39’刻伤叶片存活率为（50.00 ± 7.07）%，分化系数为（2.06 ± 0.17）。（2）最佳预培养时间为6 d，并施以80 mg/L秋水仙素浓度处理48 h，获得了纯合四倍体‘京枣39’。（3）最佳增殖培养基为MS + 0.8 mg/L 6-BA + 0.4 mg/L IBA + 30 g/L麦芽糖，pH = 5.8，平均增殖系数为（4.22 ± 0.22）。（4）最佳生根培养基为改良1/2MS + 0.8 mg/L IBA + 30 g/L麦芽糖，pH = 5.8，平均根长为（5.22 ± 0.19） cm；平均生根数为（3.20 ± 0.22）。
结论
初步建立了‘京枣39’的离体多倍体诱导体系，获得了‘京枣39’的多倍体植株，并优化了其增殖培养及生根培养方法。本研究为优良枣树品种‘京枣39’的种质创新提供了切实可行的技术方法，为多倍体诱导在枣树以及其他果树中的应用奠定了试验基础。
- ‘京枣39’ /
- 多倍体诱导 /
- 秋水仙素 /
- 增殖培养 /
- 生根培养 /
- 体系优化
Abstract:
Objective
This paper studies the effects of colchicine solution mass concentration, treatment time and pre-culture time on the induction of polyploidof ‘Jingzao 39’ in vitro, optimizes the proliferation and rooting medium of ‘Jingzao 39’, and an in vitro polyploid induction system for ‘Jingzao 39’ was established and polyploid plants were obtained.
Method
Using randomized complete block design, we investigated the effects of different mass concentrations of colchicine solution and treatment time combinations on adventitious bud regeneration of ‘Jingzao 39’, the effects of different pre-culture time on polyploid induction of ‘Jingzao 39’ in vitro, the effects of different growth regulator combinations on the proliferation culture of ‘Jingzao 39’ and the effects of different basic media and growth regulator combinations on the rooting culture of ‘Jingzao 39’.
Result
(1) When the mass concentration of colchicine solution was 80 mg/L and treated for 48 h, the survival rate of ‘Jingzao 39’ leaves was (50.00 ± 7.07)%, and the differentiation coefficient was (2.06 ± 0.17). (2) The best pre-culture time was 6 d, and the tetraploid ‘Jingzao 39’ was obtained with 80 mg/L colchicine and treatment for 48 h. (3) The optimal proliferation medium was MS + 0.8 mg/L 6-BA + 0.4 mg/L IBA + 30 g/L maltose, pH = 5.8, and the average proliferation coefficient was (4.22 ± 0.22). (4) The best rooting medium was modified 1/2MS + 0.8 mg/L IBA + 30 g/L maltose, pH = 5.8, and the average root length was (5.22 ± 0.19) cm, and the average rooting number was (3.20 ± 0.22).
Conclusion
The polyploid induction system of ‘Jingzao 39’ is established preliminarily, and one polyploid ‘Jingzao 39’ is obtained. The methods of its proliferation and rooting culture are optimized. It provides a feasible technical method for the germplasm innovation of fine jujube variety ‘Jingzao 39’, and provides an experimental basis for the application of polyploid induction in jujube and other fruit trees.
- ‘Jingzao 39’ /
- polyploid induction /
- colchicine /
- proliferation culture /
- rooting culture /
- system optimization

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图 1 不同秋水仙素质量浓度和处理时间组合对‘京枣39’不定芽存活率的影响

不同小写字母表示差异显著（P < 0.05）。下同。Different lowercase letters indicate significant differences (P < 0.05). The same below.

Figure 1. Effects of different colchicine mass concentrations and co-culture time combinations on the survival rate of adventitious buds of ‘Jingzao 39’

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图 2 不同秋水仙素质量浓度和处理时间组合对‘京枣39’不定芽分化系数的影响

Figure 2. Effects of different colchicine mass concentrations and co-culture time combinations on differentiation coefficient of adventitious buds of ‘Jingzao 39’

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图 3 80 mg/L秋水仙素溶液中处理48 h后光培养45 d‘京枣39’不定芽分化情况

Figure 3. Adventitious bud differentiation of ‘Jingzao 39’ after 45 d of photo-culture under 48 h 80 mg/L colchicine solution treating

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图 4 不同6-BA和IBA质量浓度组合对‘京枣39’增殖倍数的影响

Figure 4. Effects of different 6-BA and IBA mass concentration combinations on multiplication time of ‘Jingzao 39’

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图 5 不同6-BA和NAA质量浓度组合对‘京枣39’增殖倍数的影响

Figure 5. Effects of different 6-BA and NAA mass concentration combinations on multiplication time of ‘Jingzao 39’

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图 6 ‘京枣39’增殖培养30 d的增殖效果

Figure 6. Proliferation effects of ‘Jingzao 39’ after 30 d of proliferation culture

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图 7 不同基本培养基类型对‘京枣39’根长及根数量的影响

Figure 7. Effects of different basic medium types on rooting length and rooting number of ‘Jingzao 39’

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图 8 不同IBA质量浓度对‘京枣39’根长和根数量的影响

Figure 8. Effects of different IBA mass concentrations on root length and rooting number of ‘Jingzao 39’

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图 9 ‘京枣39’生根培养45 d的生根效果

Figure 9. Rooting effects of 45 d rooting culture of ‘Jingzao 39’

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表 1 ‘京枣39’秋水仙素处理组合

Table 1 Combination of ‘Jingzao 39’ colchicine treatment

水平 Level	因素 Factor
水平 Level	秋水仙素质量浓度 Colchicine mass concentration/（mg·L⁻¹）	处理时间 Processing time/h
1	40	48
2	60	72
3	80

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表 2 ‘京枣39’增殖培养中不同质量浓度的6-BA和IBA处理组合

Table 2 Treatment combinations of different mass concentrations of 6-BA and IBA in ‘Jingzao 39’ multiplication culture

水平 Level	因素 Factor
水平 Level	6-BA/（mg·L⁻¹）	IBA/（mg·L⁻¹）
1	1.0	0.5
2	0.8	0.4
3	0.6	0.3
4	0.4	0.2

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表 3 ‘京枣39’增殖培养中不同质量浓度的6-BA和NAA处理组合

Table 3 Treatment combinations of different mass concentrations of 6-BA and NAA in ‘Jingzao 39’ multiplication culture

水平 Level	因素 Factor
水平 Level	6-BA/（mg·L⁻¹）	NAA/（mg·L⁻¹）
1	1.0	0.5
2	0.8	0.4
3	0.6	0.3
4	0.4	0.2

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表 4 ‘京枣39’生根培养处理组合

Table 4 Treatment combinations of rooting culture of ‘Jingzao 39’

水平 Level	因素 Factor
水平 Level	基本培养基 Basic medium	IBA/（mg·L⁻¹）	6-BA/（mg·L⁻¹）
1	1/2MS	0.8	0.2
2	改良1/2MS Modified 1/2MS	0.6	0.0
3		0.4
4		0.2
注：改良1/2MS培养基参考自文献[21]。Note: modified 1/2MS medium referencd from reference [21].

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表 5 不同预培养时间‘京枣39’多倍体诱导结果

Table 5 Polyploid induction results of ‘Jingzao 39’ under different pre-culture time

预培养天数 Pre-culture day	四倍体数 Number of tetraploids	混倍体数 Number of mixoploid
4	0	0
5	0	0
6	1	0
7	0	3

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表 6 培养基组合对‘京枣39’生根情况的影响

Table 6 Effects of culture medium combinations on rooting of ‘Jingzao 39’

处理 Treatment	基本培养基 Basic medium	IBA/（mg·L⁻¹）	6-BA/（mg·L⁻¹）	根长 Root length/cm	生根量 Rooting number
1	1/2MS	0.8	0.0	3.10 ± 0.19f	1.30 ± 0.14fg
2		0.8	0.2	3.46 ± 0.21de	1.46 ± 0.15f
3		0.6	0.0	1.44 ± 0.19i	1.34 ± 0.21fg
4		0.6	0.2	1.72 ± 0.33h	1.32 ± 0.08fg
5		0.4	0.0	0.78 ± 0.15g	1.28 ± 0.13fg
6		0.4	0.2	0.96 ± 0.23g	1.16 ± 0.11g
7		0.2	0.0	0.42 ± 0.19k	1.44 ± 0.34f
8		0.2	0.2	0.46 ± 0.21k	0.96 ± 0.19h
9	改良1/2MS Modified 1/2MS	0.8	0.0	5.22 ± 0.19a	3.20 ± 0.22b
10		0.8	0.2	4.56 ± 0.17b	3.54 ± 0.15a
11		0.6	0.0	4.76 ± 0.11b	2.90 ± 0.14c
12		0.6	0.2	4.06 ± 0.11c	3.22 ± 0.08b
13		0.4	0.0	3.54 ± 0.23d	2.62 ± 0.13d
14		0.4	0.2	3.22 ± 0.19ef	2.64 ± 0.17d
15		0.2	0.0	1.30 ± 0.16i	2.44 ± 0.17de
16		0.2	0.2	2.40 ± 0.32g	2.40 ± 0.16e

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表 7 培养基组合对‘京枣39’生根长度和数量的方差分析

Table 7 Variance analysis of medium combination on rooting length and number of ‘Jingzao 39’

变异来源 Source of variation	自由度 Degree of freedom (df)	均方Mean square (MS)		F
变异来源 Source of variation	自由度 Degree of freedom (df)	根长 Root length	生根量 Rooting number	根长 Root length	生根量 Rooting number
区组 Block	4	0.056	0.018
基本培养基 Basic medium (A)	1	87.362	50.403	2 028.726*	2 139.125*
IBA (B)	3	31.355	1.314	728.132*	55.772*
6-BA (C)	1	0.025	0.010	0.569	0.430
A × B	3	2.361	0.581	54.835*	24.677*
A × C	1	0.648	0.378	15.048*	16.048*
B × C	3	0.652	0.255	15.152*	10.828*
A × B × C	3	1.190	0.024	27.634*	1.038
随机误差 Random error	60	0.042	0.024
总计 Total	79
注：表示具有显著性差异（P < 0.05）。Note: indicates significant differences (P < 0.05).

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