‘凤丹’牡丹幼胚愈伤组织诱导的优化和再生体系的建立

doi:10.16420/j.issn.0513-353x.2020-0243

摘要/Abstract

摘要：

以‘凤丹’牡丹的幼胚为材料,从取样时期、切割接种方式、基本培养基和植物生长调节剂选择等方面进行再生体系优化。结果表明：授粉后60 d为‘凤丹’牡丹幼胚诱导愈伤的最佳取材时期;完全破碎切割成幼胚薄片比完整胚、1/2胚和1/4胚更容易获得高质量的愈伤组织,且褐化率较低;MS基本培养基添加0.162 μmol · L^-1 H₂O₂可提高幼胚愈伤组织诱导率,但会增加褐化率,而在WPM基本培养基中添加不同浓度的H₂O₂,对愈伤诱导率和褐化率的影响不大;WPM基本培养基比MS更适于‘凤丹’牡丹幼胚再生;WPM + 2.5 mg · L^-1 2,4-D + 0.1 mg · L^-1 6-BA是诱导愈伤的最佳植物生长调节剂组合,诱导率可达93.33%;不定芽和不定根诱导的最佳组合分别为WPM + 30 g · L^-1蔗糖 + 3 g · L^-1植物凝胶 + 0.5 mg · L^-1 6-BA + 0.5 mg · L^-1 GA₃和1/2 WPM + 30 g · L^-1 蔗糖 + 2.5 g · L^-1植物凝胶 + 2 mg · L^-1 IBA + 1 mg · L^-1 IAA。

关键词: 牡丹, 幼胚, 愈伤诱导, 褐化, 离体, 再生

Abstract:

In order to improve callus induction quality,and establish highly efficient regeneration system of Paeonia ostii‘Fengdan’,the immature embryo collecting time,technical treatments of immature embryos,basic medium selection,and different concentrations and combinations of plant growth regulators and H₂O₂ were optimized. The results showed that the best collection time for immature embryos callus induction was 60 days after pollination. Compared with four technical treatments of immature embryos,thoroughly divided embryos（slices）were produced better callus with low callus browning rate than that with whole embryos,halved embryos,and broken into quarters embryos. MS basic medium with 0.162 μmol · L^-1 H₂O₂ can improve the callus induction rate of immature embryo,and it can also increases the callus browning rate. Different concentration of H₂O₂ adding in WPM basic medium could not be improved the callus induction rate. WPM basic medium is better than MS basic medium for immature embryo callus induction. The combination of WPM + 2.5 mg · L^-1 2,4-D + 0.1 mg · L^-1 6-BA can obtain higher callus induction rate（93.33% ± 3.53%）with lower callus browning rate（25.33% ± 4.81%）. The best hormone combinations for adventitious bud and root induction were WPM + 0.5 mg · L^-1 GA₃ + 0.5 mg · L^-1 6-BA,1/2 WPM + 2.0 mg · L^-1 IBA + 1.0 mg · L^-1 IAA,respectively.

Key words: tree peony, immature embryo, callus induction, callus browning, in vitro, regeneration

中图分类号:

S685.11

刘蓉, 慈惠婷, 任秀霞, 高洁, 王顺利, 张秀新. ‘凤丹’牡丹幼胚愈伤组织诱导的优化和再生体系的建立[J]. 园艺学报, 2022, 49(1): 166-174.

LIU Rong, CI Huiting, REN Xiuxia, GAO Jie, WANG Shunli, ZHANG Xiuxin. Optimization of Callus Induction from Immature Embryo and Establishment Regeneration System of Paeonia ostii‘Fengdan’[J]. Acta Horticulturae Sinica, 2022, 49(1): 166-174.

图/表 6

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取材时间 Embryo collection time	愈伤组织诱导率/% Callus induction rate	褐化率/% Browning rate
授粉后50 d 50 days after pollination	86.67 ± 2.81 B	4.00 ± 2.31 C
授粉后60 d 60 days after pollination	97.33 ± 2.67 A	4.00 ± 0.00 C
授粉后70 d 70 days after pollination	90.67 ± 2.67 C	21.33 ± 3.53 B
授粉后80 d 80 days after pollination	66.67 ± 3.53 D	58.67 ± 1.33 A

取材时间 Embryo collection time	愈伤组织诱导率/% Callus induction rate	褐化率/% Browning rate
授粉后50 d 50 days after pollination	86.67 ± 2.81 B	4.00 ± 2.31 C
授粉后60 d 60 days after pollination	97.33 ± 2.67 A	4.00 ± 0.00 C
授粉后70 d 70 days after pollination	90.67 ± 2.67 C	21.33 ± 3.53 B
授粉后80 d 80 days after pollination	66.67 ± 3.53 D	58.67 ± 1.33 A

接种方法 Inoculation method	愈伤诱导率/% Callus induction rate	褐化率/% Browning rate	愈伤组织特征 Characteristics of callus
完整胚The intact embryo	100.00	60.00 ± 1.67 a	白色,较松软 White,soft
1/2胚 The 1/2 embryo	100.00	45.00 ± 3.33 b	白色,表面有雪花和松散的结构 White,with snowflakes and loose structure on the surface
1/4胚 The 1/4 embryo	100.00	25.00 ± 1.67 c	淡黄色,结构紧密 Light yellow,compact structure
幼胚薄片 The young embryo slice	100.00	15.00 ± 1.67 d	淡黄色,结构紧密,颗粒状 Light yellow,compact structure,granular

接种方法 Inoculation method	愈伤诱导率/% Callus induction rate	褐化率/% Browning rate	愈伤组织特征 Characteristics of callus
完整胚The intact embryo	100.00	60.00 ± 1.67 a	白色,较松软 White,soft
1/2胚 The 1/2 embryo	100.00	45.00 ± 3.33 b	白色,表面有雪花和松散的结构 White,with snowflakes and loose structure on the surface
1/4胚 The 1/4 embryo	100.00	25.00 ± 1.67 c	淡黄色,结构紧密 Light yellow,compact structure
幼胚薄片 The young embryo slice	100.00	15.00 ± 1.67 d	淡黄色,结构紧密,颗粒状 Light yellow,compact structure,granular

培养基 Culture media	H₂O₂/（μmol · L^-1）	愈伤组织诱导率/% Callus induction rate	愈伤组织褐化率/% Browning rate of callus
MS	0（对照 Control）	66.67 ± 10.90 b	28.00 ± 2.31 b
	0.162	90.67 ± 3.53 a	41.33 ± 13.33 a
	0.324	84.00 ± 2.31 ab	49.33 ± 12.72 a
	0.486	72.00 ± 8.33 b	29.33 ± 2.31 b
WPM	0（对照 Control）	97.33 ± 2.67 a	4.00 ± 0.00 c
	0.162	97.33 ± 1.33 a	4.00 ± 2.31 c
	0.324	92.00 ± 4.00 a	2.67 ± 1.33 c
	0.486	92.00 ± 4.62 a	2.67 ± 2.67 c