月季红双喜花瓣变色的化学基础及比较转录组分析

doi:10.11983/CBB22049

摘要/Abstract

摘要： 月季(Rosa hybrida)花色丰富, 是世界著名的观赏花卉。月季红双喜因其花瓣的变色特性而在市场上广受欢迎。该研究通过类黄酮和类胡萝卜素靶向代谢并结合转录组分析, 发现红双喜的黄色花瓣呈色主要源于叶绿素、类胡萝卜素以及类黄酮的积累, 红色花瓣呈色主要是花青素苷积累增加且糖苷化的结果。花青素苷合成关键基因CHI、ANS和UFGT, 以及R2R3-MYB家族的AN2-like成员在红色花瓣中的强烈表达是花青素苷积累的分子基础; 类胡萝卜素成分改变及相关基因的表达变化在红双喜花瓣变色过程中也起重要作用, 并且miRNA156可能参与其调控过程。该研究揭示了月季红双喜花瓣变色的分子和化学基础, 研究结果为观赏植物花色分子设计育种提供了重要理论依据。

关键词: 月季, 花色, 花青素苷, 类胡萝卜素, 分子调控

Abstract: Rosa species is one of world-famous ornamental flower with very rich colors. Rosa hybrida cv. ‘Double delight’ is very popular in the market due to the color-changing properties of petals. With analysis of the composition of flavonoids and carotenoids, combined with transcriptome analysis, we showed that the red petals were caused by the increasing of glycosylated anthocyanins, and the up-regulation of anthocyanin synthesis genes CHI, ANS and UFGT by R2R3-MYB transcription factor AN2-likes. However, the yellow petals were mainly due to the accumulation of the pigments of chlorophyll, carotenoids and flavonoids. The structural differences of carotenoids and the expression changes of related genes may play an important role in the petals’ discoloration, and miRNA156 might be involved in the regulation. This study revealed a molecular and chemical basis of the petal discoloration in rose Double delight and provided an important basis for the molecule design and breeding of flowers.

Key words: rose, flower color, anthocyanins, carotenoids, molecular regulation

张和臣, 王慧娟, 李艳敏, 高杰, 袁欣, 王利民, 王校晨, 赵银鸽, 符真珠. 月季红双喜花瓣变色的化学基础及比较转录组分析. 植物学报, 2022, 57(5): 649-660.

Zhang Hechen, Wang Huijuan, Li Yanmin, Gao Jie, Yuan Xin, Wang Limin, Wang Xiaochen, Zhao Yinge, Fu Zhenzhu. The Chemical Composition and Transcriptome Analysis Reveal the Mechanism of Color Formation in Rosa hybrida cv. ‘Double delight’. Chinese Bulletin of Botany, 2022, 57(5): 649-660.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB22049

https://www.chinbullbotany.com/CN/Y2022/V57/I5/649

图/表 10

表1 qRT-PCR所用引物

Table 1 The primers used for qRT-PCR

Gene name	Pri- mers	Sequences (5′-3′)
CHI	F	GCAATACTCGGAGAAGGTTTCA
(LOC112182551)	R	CAATCACCGCATTTCCAAC
ANS	F	TAGAAGAAGGGAGGCTGGAG
(LOC112179310)	R	TGTGGAGGATGAAGGTGAGT
UFGT	F	TTGTAACACACTGCGGGTG
(LOC112172868)	R	GAACATCTCTGAGCATTCGTG
PSY	F	GCTGTTGCTCACCCATCAAG
(LOC112190337)	R	CAAACCTCACCACACCTATCG
LYCB	F	ACACAGACCCTTCCCTCCAA
(LOC112188432)	R	TGGTTCTTCCACAACGGTTT
ZDE	F	CCTGCCTGTCAATCTTGTAGAC
(LOC112189356)	R	TCCCACTATCACCACATCCTC
AN2-like1	F	GCTGTAGACTGAGGTGGCTAAA
(LOC112185634)	R	GTGAAAGGACGATGGGCTA
AN2-like2	F	GGACGAACTGGAAACGATG
(LOC112193894)	R	GTGATGCTTGTGTTGAGCG
AN2-like3	F	GGAAGATGGCACAAGGTTC
(LOC112186121)	R	GCCGAGCACTCCAATAGTTT
UVR8	F	GGCAGAGTTCTTTCTTGACAGAC
(LOC112182836)	R	GGCAATGCTGAGAGAGTTTCA
PIF3	F	TGATGAGAAGATTGACCGAGG
(LOC112192990)	R	AGAAGACGGCGAAAGGCTA
HY5	F	GGCATACTTGAGTGACTTGGAA
(LOC112172411)	R	CGGCTTGCTGTTGTGTTCT
GAPDH	F	TATGACCAGATCAAGGCTGCT
(JN399220)	R	ACCAATGAAGTCGGTTGACAC

表2 月季红双喜不同呈色花瓣中主要类胡萝卜素的成分含量差异

Table 2 Differential carotenoid components in different colored petals of Rosa hybrida cv. ‘Double delight’

No.	Type of carotenoid	Y1 (μg·g^-1)	Y2 (μg·g^-1)	R1 (μg·g^-1)	R2 (μg·g^-1)	Log₂FC	Regulated
Carotenoid_22	Neochrome palmitate	2.2016998	2.3834704	1.0197834	1.0475690	-1.1491910	Down
Carotenoid_24	Rubixanthin laurate	0.1569250	0.1481073	0.0527210	0.0507537	-1.5596845	Down
Carotenoid_32	Violaxanthin dilaurate	3.9240426	4.7154605	2.9313100	2.8637581	-0.5761227	Unchanged
Carotenoid_33	Violaxanthin-myristate-caprate	21.7732951	22.4958882	12.8019824	12.7492289	-0.7929111	Unchanged
Carotenoid_34	Violaxanthin-myristate-laurate	5.6980954	6.7643503	4.7010832	5.0801542	-0.3494983	Unchanged
Carotenoid_39	Violaxanthin dioleate	0.7922281	0.8738322	0.4032495	0.3662500	-1.11444831	Down
Carotenoid_41	Zeaxanthin palmitate	0.1189156	0.1116267	0.3534273	0.3530580	1.61562841	Up
Carotenoid_51	β-cryptoxanthin laurate	0.7270018	0.7903577	0.3395545	0.3316843	-1.1766652	Down
Carotenoid_54	β-cryptoxanthin oleate	0.1793999	0.1468413	0.0389599	0.0467119	-1.9290470	Down
Carotenoid_56	Zeaxanthin	5.9183084	6.1692434	17.8138771	17.0472808	1.5280984	Up
Carotenoid_57	Violaxanthin	4.9150727	4.8171053	3.8840384	3.8675731	-0.3282664	Unchanged
Carotenoid_58	Neoxanthin	2.0484948	2.2917558	1.9106499	1.9435004	-0.1713655	Unchanged
Carotenoid_59	Lutein	7.8872005	8.3871299	18.9305539	18.6268263	1.2064982	Up
Carotenoid_66	Canthaxanthin	0.0005338	0.0004385	0	0	-Inf	Down

图1 月季红双喜不同发育时期花瓣呈色表型(A)、不同呈色花瓣类型中的类黄酮差异(B)及主要差异成分(C) S1: 萼片初展期; S2: 花瓣透色期; S3: 花瓣初开期; S4: 花瓣盛开期

Figure 1 The phenotype of the petal coloration (A), the difference of flavonoids (B), and the main different components (C) in petals of Rosa hybrida cv. ‘Double delight’ S1: The development stage when sepals are initially unfolded; S2: The development stage when petals are being colored; S3: The development stage when petals are just opened; S4: The development stage when petals are in full bloom

表3 月季红双喜不同呈色花瓣中mRNA、miRNA和LncRNA的数量

Table 3 Transcripts of mRNA, miRNA and LncRNA in different colored petals of Rosa hybrida cv. ‘Double delight’

Type	Total transcripts	Differentially expressed transcripts	Up regu- lated	Down regu- lated
mRNA	36193	2250	1371	879
miRNA	7845	22	21	1
LncRNA	146605	51	24	27

表4 月季红双喜不同呈色花瓣中差异表达基因的功能注释

Table 4 Functional annotation of differentially expressed genes in different colored petals of Rosa hybrida cv. ‘Double delight’

Comparison set	Total DEG	COG	GO	KEGG	KOG	NR	Pfam	Swiss-Prot	eggNOG
Y_vs_R	2177	965	1904	1596	1034	2177	1923	1733	1858

图2 月季红双喜不同呈色花瓣中差异LncRNA靶基因聚类(A)及差异siRNA靶基因聚类(B)

Figure 2 Analysis of differential LncRNA target genes (A) and siRNA target genes (B) clustering in different colored petals of Rosa hybrida cv. ‘Double delight’

图3 月季红双喜不同呈色花瓣中的基因转录特性(A)及差异表达基因KEGG聚类(B) FDR: 错误发现率

Figure 3 Characteristics of gene expression (A) and KEGG clustering of DEGs (B) in different colored petals of Rosa hybrida cv. ‘Double delight’ FDR: False discovery rate

图4 月季红双喜花瓣中花青素苷及类胡萝卜素合成通路相关基因的表达特性分析蓝色至红色代表基因表达由弱至强(彩图见网站版本)。*代表差异显著基因。

Figure 4 Expression pattern analysis of anthocyanin and carotenoid synthesis-related genes in the petals of Rosa hybrida cv. ‘Double delight’ The blue to red color represents gene expression from weak to strong (the color version is shown in online). *represent genes with significant difference.

图5 月季红双喜不同呈色花瓣中差异表达R2R3-MYB基因和类黄酮调控相关R2R3-MYB基因进化分析(A)及R2R3-MYB相关基因的表达热图(B) Y和R同表2。*代表与花青素苷合成调控相关的R2R3-MYB基因。

Figure 5 Phylogenic analysis of differentially expressed R2R3-MYB and flavonoid regulation related genes (A), and heat map of the corresponding R2R3-MYB members in different colored petals of Rosa hybrida cv. ‘Double delight’ (B) Y and R see Table 2. * represent R2R3-MYB genes related to the regulation of anthocyanin biosynthesis.

图6 qRT-PCR验证相关基因在月季红双喜花瓣不同发育时期的表达特性 S1-S4同图1。*表示与S1时期相比差异显著。

Figure 6 qRT-PCR analysis of the expression pattern of the corresponding genes in the Rosa hybrida cv. ‘Double delight’ petals at different developmental stages S1-S4 see Figure 1. * mean significant differences compared with S1 stage.

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