林兵^,1, 陈艺荃², 钟淮钦¹, 叶秀仙¹, 樊荣辉^,11福建省农业科学院作物研究所,福州 350013
²福建省农业科学院农业工程技术研究所,福州 350013

Analysis of Key Genes About Flower Color Variation in Iris hollandica

LIN Bing^,1, CHEN YiQuan², ZHONG HuaiQin¹, YE XiuXian¹, FAN RongHui^,11Institute of Crop Sciences, Fujian Academy of Agricultural Science, Fuzhou 350013
²Institute of Agriculture and Engineering Technology, Fujian Academy of Agricultural Science, Fuzhou 350013

通讯作者: 樊荣辉,E-mail：rhfan1012@163.com

责任编辑: 赵伶俐
收稿日期:2020-08-24接受日期:2020-12-8网络出版日期:2021-06-16

基金资助:

福建省省属公益类科研专项.2018R1026-6 福建省农业科学院科技创新团队项目.STIT2017-2-9 福建省自然科学基金.2018J01045

Received:2020-08-24Accepted:2020-12-8Online:2021-06-16
作者简介 About authors
林兵,E-mail：lb87572540@163.com。







摘要
【目的】花色变异对丰富观赏植物花色具有重要意义,但由于花色变异的不确定性,使其变异机理尚未弄清。荷兰鸢尾是重要球根观赏植物,通过探索荷兰鸢尾蓝紫色野生型‘展翅’及白色突变株‘玉妃’的显色分子机制及色素积累差异,为花色变异机理提供依据。【方法】本研究通过超高效液相色谱-四级杆飞行时间串联质谱联用（UHPLC-QTOF-MS）方法测定荷兰鸢尾2个品种花的花色素苷和黄酮醇的种类及含量。以荷兰鸢尾‘展翅’和‘玉妃’的旗瓣为材料,通过转录组测序,筛选花色素苷合成相关差异基因。以2个品种花发育不同时期为材料,对差异基因进行荧光定量PCR验证。【结果】代谢组分析结果表明,飞燕草素和矢车菊素及其衍生物在蓝紫色花中积累,在白色花中几乎没有积累,而黄酮醇类物质在白色‘玉妃’中含量上升。RNA-seq结果表明,共获得46 485个unigenes,其中27 073个unigenes被公共数据库功能注释,占全部unigenes的58.24%。获得701个差异表达基因,其中485个基因在‘玉妃’中上调表达,216个基因在‘玉妃’中下调表达。花色素苷途径中,2个二氢黄酮醇-4-还原酶（dihydroflavonol 4-reductase,DFR）基因和1个黄酮醇合成酶（flavonol synthase,FLS）基因有差异表达,分别命名为IhDFR1、IhDFR2和IhFLS1。白色‘玉妃’中,IhDFR1和IhDFR2下调表达,IhFLS1上调表达,导致花色素苷含量显著降低和黄酮醇积累,使代谢流由花色素苷转向黄酮醇。3个基因的qRT-PCR结果显示,IhDFR1和 IhDFR2在蓝紫色花中随着花的发育表达量升高,在白色花中均低表达,IhFLS1在白色花中随着花的发育表达量升高,在蓝紫色花中均低表达,与RNA-seq结果保持一致。【结论】白色‘玉妃’中,IhDFR1和IhDFR2的低表达,及IhFLS1的高表达,使花色素苷积累受阻,部分代谢流由花色素苷转向黄酮醇,导致花色由蓝紫变为白色。
关键词： 荷兰鸢尾;花色素苷;黄酮醇;转录组分析;二氢黄酮醇-4-还原酶基因;黄酮醇合成酶基因

Abstract
【Objective】 Flower color variation is of great significance for enriching color of ornamental plants, but it is difficult to clarify variation mechanism due to uncertainty of flower color variation. Dutch iris (Iris hollandica) is an important bulbous ornamental plant. In this study, the blue-purple wild type ‘Zhanchi’ and white mutant strain 'Yufei' of Dutch iris were investigated to explore molecular mechanism and difference of pigment accumulation, so as to provide a basis for mechanism of flower color variation.【Method】In this study, using inner tepals of ‘Zhanchi’ and ‘Yufei’ from Dutch Iris as materials, UHPLC-QTOF-MS method was used to determine types and contents of anthocyanins and flavonols from two varieties of Dutch Iris, and the different expression genes related to anthocyanin synthesis were screened by transcriptome sequencing. Using different flowers in developmental stages of two varieties as materials, the different expression genes were verified by qRT-PCR.【Result】Results of metabolomics analysis revealed that delphinidin, cyanidin and its derivatives were accumulated in blue-purple flowers, which were almost no accumulation in white flowers, while the flavonol contents were increased in white ‘Yufei’. Results of RNA-seq analysis revealed that a total of 46 485 unigenes were obtained, and 27 073 unigenes of them were functionally annotated by public databases, accounting for 41.85% of the total. And 701 differentially expressed genes were obtained, 485 genes of which were up-regulated and 216 genes were down-regulated in white ‘Yufei’. Two dihydroflavonol-4-reductase genes and one flavonol synthase gene involving in anthocyanin biosynthetic pathway had different expression, named IhDFR1, IhDFR2 and IhFLS1. Down-regulated expression of IhDFR1 and IhDFR2 as well as up-regulated expression of IhFLS1 in white ‘Yufei’ led to significant decrease of anthocyanins and accumulation of flavonols, which caused metabolic flow from anthocyanin to flavonol. The qRT-PCR results of three genes showed that expression levels of IhDFR1 and IhDFR2 increased in blue-purple flowers during flower developmental stage, but low expression in white flowers, and expression level of IhFLS1 increased in white flowers during flower developmental stage, but low expression in blue-purple flowers, which was consistent with RNA-seq results.【Conclusion】Low expression of IhDFR1 and IhDFR2 as well as high expression of IhFLS1 in white ‘Yufei’ blocked accumulation of anthocyanins, and some of metabolic flow changed from anthocyanins to flavonols, resulting in the change of flower color from blue violet to white.
Keywords：Iris hollandica;anthocyanin;flavonol;transcriptome analyses;dihydroflavonol-4-reductase gene;flavonol synthase gene


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本文引用格式
林兵, 陈艺荃, 钟淮钦, 叶秀仙, 樊荣辉. 荷兰鸢尾‘玉妃’花色变异关键结构基因分析[J]. 中国农业科学, 2021, 54(12): 2644-2652 doi:10.3864/j.issn.0578-1752.2021.12.014
LIN Bing, CHEN YiQuan, ZHONG HuaiQin, YE XiuXian, FAN RongHui. Analysis of Key Genes About Flower Color Variation in Iris hollandica[J]. Scientia Acricultura Sinica, 2021, 54(12): 2644-2652 doi:10.3864/j.issn.0578-1752.2021.12.014


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0 引言

【研究意义】花色是观赏植物重要的性状特征^[1],花色变异对丰富其花色具有重要意义,阐明花色变异机理,可为花色定向改良提供依据。【前人研究进展】荷兰鸢尾（Iris hollandica）为鸢尾科（Iridaceae）鸢尾属（Iris spp.）秋植球根花卉。花姿优美、如鸢似蝶、花色艳丽,主要有白色、黄色和蓝紫色等色系^[2]。许多植物中,由红到紫到蓝色花的形成主要由花色素苷途径控制,如茶树（Camellia sinensis (L.)）、葡萄风信子（Muscari）、睡莲（Nymphaea spp.）等^[3,4,5]。花色素苷途径在F3H、F3'H和F3'5'H三个关键酶作用下,形成3个分支,形成3种不同的二氢黄酮醇,DFR选择性地催化3种二氢黄酮醇,形成相应的花青素苷。因此,3个羟化酶F3H、F3'H、F3'5'H活性的有无、强弱、DFR的底物催化选择性及FLS对底物的竞争性是决定花色的主要因素^[6,7]。该途径结构基因的突变可能导致其编码的酶失活进而使植物中花色素苷含量降低,表型上会表现出颜色变浅、变白等特征^[8,9]。荷兰鸢尾蓝紫色花的深浅主要由花色素苷的种类和含量、黄酮助色素的辅助着色及类胡萝卜素的含量三者共同作用,其中花色素苷的种类和含量为主控因素^[10],且荷兰鸢尾花色素苷途径大部分相关酶和基因被挖掘和研究^[11,12,13],这为花色研究奠定了基础。【本研究切入点】由于特定位点突变导致色素缺失,进而导致花色变浅或变白的可能分子机制存在广泛性和不确定性,例如,可能是某个基因、某个蛋白或某个转录因子的突变,使花色变浅或变白机理的研究难以澄清,这就需要更多的数据去解释花色变异原因。【拟解决的关键问题】本研究通过探索荷兰鸢尾蓝紫色野生型‘展翅’及白色突变株‘玉妃’的显色分子机制及色素积累差异,为花色变异机理研究提供技术支持,同时对荷兰鸢尾白色花和蓝紫色花中花色苷积累机制提供更全面的了解。

1 材料与方法

1.1 试验材料

野生型‘展翅’（蓝紫色）和突变株‘玉妃’（白色）种植于福建省农业科学院花卉研究中心种质资源圃中。

2019年3月,分别取‘展翅’（蓝紫色）和‘玉妃’（白色）旗瓣,液氮中速冻,-80℃保存,用于色素成分分析、转录组测序;分别取‘展翅’和‘玉妃’花发育3个时期（花蕾前期：蕾为白色,未露出花被;花蕾中期：蕾略显色,尖端露出花被;始花期：花苞半开放）,用于qRT-PCR,以检测目的基因随花发育的表达情况。

1.2 类黄酮的定性定量分析

类黄酮的定量定性分析采用超高效液相色谱—四级杆飞行时间串联质谱联用（UHPLC-QTOF-MS）技术进行,每个样品3个生物学重复。

类黄酮的提取：冻干机中冻干后,取出约100 mg,采用上海净信实业发展有限公司型号为JX-24的全自动组织研磨仪,于40 Hz频率研磨4 min;加入1 mL 60%乙醇溶液（含0.1%盐酸）于恒温水浴锅中35℃下浸提2 h;在4℃离心机中14 000 r/min离心15 min,取上清,用温和氮气吹去其中的乙醇,并补充一定体积0.1%盐酸的水溶液,用于后续分析。

UPLC条件：色谱柱为ACQUITY UPLC BEH C18（100 mm×2.1 mm,1.7 μm,Waters）,温度为45℃。流动相组：A相,水（含0.5%甲酸溶液）;B相,乙腈（含0.5%甲酸溶液）。洗脱条件：2 min,1% B;~1 min,5% B;6 min,20% B;3 min,50% B;3 min,100% B;100% B保持2 min;最后1% B平衡3 min。流速0.4 mL·min^-1,进样体积3 μL。

质谱条件：LockSpray离子源在正电喷雾电离和负电喷雾电离（ESI）模式下运行。扫描模式为MSE模式,以低能扫描（CE 4 eV）和高能扫描（CE倾斜20—45 eV）来破碎离子,氩气（99.999%）作离解气体。扫描范围50—1 000 amu,速度0.2 s/扫描。毛细管电压2 kV（正模式）,锥电压40 V,源电压偏移60 V。光源温度115℃,脱溶剂气温度为450℃。去溶剂气流量900 L·h^-1,锥气流量50 L·h^-1,氮气（>99.5%）用作去溶剂和锥气。

花色素苷和黄酮醇含量测定采用标准曲线法,利用标准品矢车菊素3-O-葡萄糖苷（Cyanidin-3-O- glucoside）和槲皮素（Quercetin）作为外标进行半定量分析。标准品的浓度梯度为500、250、100、50、10、5和1 μg·mL^-1。每个样品重复3次。采用UNIFI 1.8.1（Waters）软件进行数据分析。

1.3 转录组测序及分析

通用RNA提取试剂盒（北京百泰克生物科技有限公司）提取总RNA。RNA完整性用NanoDrop 2000（Thermo Scientific,MA,USA）检查。使用Illumina HiSeq?4000进行文库构建和转录组测序（北京百迈克生物技术公司）。

转录组数据分析流程如下：应用FASTX-Toolkit软件获得原始Raw reads,利用FastQC（http://www.bioinformatics.babraham.ac.uk/projects/fastqc）软件对Raw reads过滤得到高质量的Clean reads;Clean reads再组装成Transcripts和Unigene。利用RSEM软件对基因表达量进行预测,采用FPKM（Fragments Per Kilobase Million）方法对测序数据标准化处理^[14]。使用DESeq软件对差异表达基因进行评估^[15]。差异基因间的阈值为false discovery rate（FDR）<0.01 a且fold- change value≥2。由8大数据库进行注释,包括Nr、Nt、GO、KOG、KEGG、COG、Pfam和eggNOG。

1.4 实时荧光定量PCR（qRT-PCR）分析

使用ABI 7500实时PCR系统（Applied Biosystems）进行qRT-PCR。应用Primer Premier 5.0进行引物设计以确定基因的相对表达水平。熔解曲线分析确认PCR的特异性。每个样品3个生物学重复。以β-actin为内参。相对定量计算方法应用2^-ΔΔCt法。

Table 1
表1
表1qRT-PCR所用引物序列
Table 1Primer sequences of qRT-PCR

基因 Gene	正向引物（5′-3′）Forward primer sequence	反向引物（5′-3′）Reverse primer sequence
IhDFR1	GAGGTGGTCGCAGGATGCACT	CTCCGTCTGCTGATGTTCTTT
IhDFR2	AAGGCGGTCGCAGGATGCACC	TGATGAAAGGACCGACGACT
IhFLS1	GTTGGAGTGATGGACGGGATG	GGGACAGGGAGGGTAGTAGTTGA
Ihactin	ACGGAAATTGTATGTGGGT	CAGATGCGAAAGATGTGAG

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2 结果

2.1 花色素苷及黄酮醇鉴定

应用UHPLC-QTOF-MS技术,对野生型‘展翅’（蓝紫色）和突变品种‘玉妃’（白色）的旗瓣进行花色素苷和黄酮醇定量定性鉴定。共有9种花色素苷,‘展翅’花中色素物质主要为飞燕草素苷和矢车菊素苷,使其花色呈蓝紫色,主要含量较高的有飞燕草素-3-(6-p-酰基葡萄糖苷)-5-(6-丙二酰基-4-鼠李糖基葡萄糖苷)和矢车菊素-3-芸香糖苷,含量分别为209.85和144.42 μg·g^-1,其次为矮牵牛素-3-(6-鼠李糖基-2-木糖基葡萄糖苷)和飞燕草素-3-[6-(4-咖啡酰基木糖基)葡萄糖苷]-5-葡萄糖苷,含量分别为61.86和61.26 μg·g^-1。而‘玉妃’的9种花色素苷含量相较于‘展翅’均显著降低,预测可能是花色素苷合成途径的上游基因发生了突变,阻断了整个花色素苷代谢流的通路,使显色的花色素含量达不到显色程度,导致变异品种‘玉妃’花色变为白色。

共鉴定出14种黄酮醇,其中杨梅素-3-O-葡萄糖苷和杨梅素-3-O-半乳糖苷在‘玉妃’中含量显著升高,分别为145.22和58.65 μg·g^-1,而在‘展翅’中的含量分别为3.29和4.04 μg·g^-1,预测花色素苷合成途径的部分代谢流流向黄酮醇。

2.2 转录组数据分析

为了进一步探索‘玉妃’突变为白色的原因,将‘展翅’和‘玉妃’的旗瓣进行转录组测序,共获得21.51 Gb Clean reads（表3）,通过组装获得46 485个unigenes,平均长度为839 bp,其中12 500个unigene长于1 000 bp,占总数的26.89%。‘玉妃’与‘展翅’相比,有485个基因上调表达,有216个基因下调表达。

在46 485个unigenes中,58.24%的unigenes被8大公共数据库注释。为了鉴定与次级代谢途径相关的unigenes,进行KEGG注释。约有9 909（21.31%）个unigenes分属于65个KEGG途径。类黄酮途径（花色素苷途径）是次级代谢途径中含有差异基因最多的一组之一（图2）,这与突变株‘玉妃’花色变异明显是一致的,这些基因分析为荷兰鸢尾基因挖掘和功能分析提供宝贵资源。花色素苷途径中共发现7个差异表达基因,2个二氢黄酮醇-4-还原酶（dihydroflavonol 4-reductase,DFR）基因在‘玉妃’中下调表达,1个黄酮醇合成酶（flavonol synthase,FLS）基因、1个查尔酮合成酶基因（chalcone synthase,CHS）、1个查尔酮异构酶基因（chalcone isomerase,CHI）和1个类黄酮3′-羟化酶基因（flavonoid 3′-hydroxylase,F3'H）在‘玉妃’中均上调表达。

图1

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图1荷兰鸢尾野生型‘展翅’及突变品种‘玉妃’

A：展翅 Zhanchi;B：玉妃 Yufei
Fig. 1The parent Zhanchi and the mutant variety Yufei in Iris hollandica

图2

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图2荷兰鸢尾2个品种的差异基因KEGG分析

Fig. 2KEGG analysis of DEGs involved in two varieties of Iris hollandica



Table 2
表2
表2荷兰鸢尾花中花色素苷和黄酮醇的鉴定
Table 2Identification of anthocyanin and flavonol in Iris hollandica flower

化合物种类 Species of compounds	Compound 化合物名称	含量 Content (μg·g-1)
		展翅 Zhanchi	玉妃 Yufei
花青素苷Anthocyanidin	矢车菊素-3-(2G-木糖基芸香糖苷) Cyanidin -3-(2G-xylosylrutinoside)	25.22±1.66	1.34±0.05
	飞燕草素-3-[6-(4-咖啡酰基木糖基)葡萄糖苷]-5-葡萄糖苷 Delphinidin-3-[6-(4-(caffeoylrhamnosyl) glucoside]-5-glucoside	61.26±1.33	1.36±0.08
	矢车菊素-3-芸香糖苷 Cyanidin -3-rutinoside	144.42±6.46	16.90±0.63
	6-羟基矢车菊素-3-葡萄糖苷 6-Hydroxycyanidin -3-glucoside	5.88±0.19	0.56±0.01
	飞燕草素-3-(6-p-酰基葡萄糖苷)-5-(6-丙二酰基-4-鼠李糖基葡萄糖苷) Delphinidin-3-(6-p-coumaroylglucoside)-5-[6-(malonyl)-4-(rhamnosyl) glucoside)]	209.85±3.12	1.71±0.05
	飞燕草素-3-葡萄糖苷 Delphinidin-3-glucoside	4.37±0.24	0.08±0.01
	飞燕草素-3-芸香糖苷 Delphinidin 3-rutinoside	30.64±0.38	16.98±0.48
	矮牵牛素-3-(6-鼠李糖基-2-木糖基葡萄糖苷) Petunidin-3-[6-(rhamnosyl)-2-(xylosyl) glucoside]	61.86±0.59	8.29±0.20
	总计 Total	543.5	47.22
黄酮醇 Flavonol	山奈酚-3-O-半乳糖苷 Kaempferol-3-O-galactoside	73.37±2.13	69.21±1.69
	山奈酚-7-O-葡萄糖苷 Kaempferol-7-O-glucoside	62.36±2.61	56.64±0.85
	槲皮素-3-O-葡萄糖苷 Quercetin-3-O-glucoside	21.45±0.32	12.81±0.24
	鼠李素-3-O-葡萄糖苷 Rhamnetin-3-O-Glucoside	40.01±0.51	22.95±0.32
	异鼠李素-3-O-葡萄糖苷 Isorhamnetin-3-O-Glucoside	37.02±1.20	24.14±0.67
	苜蓿素-4'-甲基醚-3'-O-葡萄糖苷 Tricin-4'-methylether-3'-O-glucoside	39.31±0.45	23.55±0.33
	杨梅素-3-O-葡萄糖苷 Myricetin-3-O-glucoside	3.29±0.08	145.22±3.74
	杨梅素-3-O-半乳糖苷 Myricetin-3-O-galactoside	4.04±0.12	58.65±2.52
	槲皮素-3-O-(6''-丙二酰)半乳糖苷 Quercetin-3-O-(6''-malonyl)galactoside	4.99±0.12	6.27±0.32
	鼠李素-3-O-芸香糖苷 Rhamnetin-3-O-Rutinoside	4.36±0.21	15.05±1.23
	3,5,7,4'-四羟基-8-甲氧基黄酮-3-O-葡萄糖苷-7-O-鼠李糖苷 Sexangularetin-3-O-glucoside-7-O-rhamnoside	4.48±0.09	14.39±0.41
	羟基山奈酚-3,6-O-二葡萄糖苷 6-Hydroxykaempferol-3,6-O-Diglucoside	4.89±0.25	1.21±0.06
	槲皮素-3-O-芸香糖苷-7-O-鼠李糖苷 Quercetin-3-O-rutinoside-7-O-rhamnoside	28.55±1.01	3.75±0.31
	槲皮素-7-O-芸香糖苷-4'-O-葡萄糖苷 Quercetin-7-O-rutinoside-4'-O-glucoside	5.04±0.06	0.02±0.01
	总计 Total	333.15	453.81

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Table 3
表3
表3转录组测序数据
Table 3RNAseq data statistics

样品 Sample	Clean read 数量 Number of clean reads	GC含量 GC content (%)	%≥Q30
展翅 Zhanchi	45850456	50.29	87.76%
玉妃 Yufei	48903721	50.30	87.08%

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2.3 花色变异相关基因分析

DFR和FLS均以二氢黄酮醇（dihydroflavonol）为底物,分别形成有色的花色素苷（anthocyanin）和无色的黄酮醇（flavonol）,两个基因间存在竞争关系。转录组数据中共发现5个DFR基因,其中3个在‘展翅’和‘玉妃’中均低表达,2个（IhDFR1和IhDFR2）在白色‘玉妃’花中表达量显著降低（图4）。共发现10个FLS基因,其中8个在‘展翅’和‘玉妃’中均低表达,1个在‘展翅’和‘玉妃’中的FPKM均为10,1个（IhFLS1）在白色‘玉妃’花中表达量显著升高。DFR聚类分析表明,IhDFR1和IhDFR2亲缘关系很近,且与同为鸢尾属的溪荪（Iris sanguinea）和马蔺（Iris lactea）聚为一类。FLS聚类分析表明,IhFLS1与其他FLS氨基酸序列亲缘关系较远,单独聚为一类（图3）。

图3

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图3荷兰鸢尾DFR及FLS进化树分析

Fig. 3Phylogenetic analysis of DFR and FLS in Iris hollandica



对‘展翅’和‘玉妃’的3个花发育时期进行qRT- PCR分析,结果显示,IhDFR1和IhDFR2在‘展翅’中随着花的发育,表达量逐渐上升,在始花期达到最高,而在‘玉妃’中花发育3个时期表达量相近。在始花期,IhDFR1和IhDFR2在‘展翅’中的表达明显高于‘玉妃’中。IhFLS1在‘展翅’中随着花的发育略有上升,至始花期达到最高,在‘玉妃’中随着花的发育显著上升,至始花期达到最高。在始花期,IhFLS1在‘玉妃’中的表达明显高于 ‘展翅’（图4）。推测在‘玉妃’中,IhDFR1和IhDFR2的低表达及IhFLS1的高表达,使花色素苷生物合成途径的花色素苷合成受阻,部分代谢流流向无色的黄酮醇,花色由蓝紫色变为白色。

3 讨论

飞燕草色素苷使花色呈现紫色到蓝色,矢车菊色素苷使花色呈现红色到品红^[16,17]。蓝紫色‘展翅’花中主要含有飞燕草素苷和矢车菊素苷,‘玉妃’花中飞燕草素苷和矢车菊素苷含量显著降低,呈现白色。然而,花色素苷生物合成途径的一些关键基因如CHS、CHI和F3'H在白色‘玉妃’花中表达量分别提高了3.2、2.2、2.3倍。前人研究中,花色素苷的积累会伴随生物合成途径中相关基因的高表达,而花色素苷的缺失会伴随相关基因的低表达^{[18,19,20,21]},本研究与前人研究存在差异。LOU等^[4]的研究中,葡萄风信子的白色花中,CHS、F3'H和F3'5'H的高表达,使该途径中不显色的黄梅酮（Myricetin）和山奈酚（Kaempferol）含量高。CHS、CHI和F3'H高表达,使白色‘玉妃’花中不显色的黄酮醇总含量升高。

DFR能催化二氢黄酮醇形成原花色素苷（Leucoanthocyanins）,进而形成花色素苷,FLS催化二氢黄酮醇形成无色的黄酮醇,与DFR存在竞争底物关系^[22,23]。由于2个基因对底物的竞争,FLS的上调表达可能会导致DFR表达量下降和花色素苷的降低。在烟草中,过表达FLS,使DFR的表达受阻,黄酮醇积累,花色变为白色^[24]。本研究白色‘玉妃’花中,DFR的低表达可能是蓝紫色色素缺失的主要原因,此外,FLS的高表达,导致其与DFR竞争二氢黄酮醇底物,部分阻止花色素苷的合成,并使代谢流流向无色的黄酮醇,最终导致蓝紫色变为白色。

目前,许多植物出现花色退化或变白现象,除DFR突变外,导致这种现象的原因有多种。如单个CHS的突变会导致花色退化,在矮牵牛（Petunia hybrida）和紫罗兰（Matthiola incana）中,由于CHS的突变,花色均变为白色^[25,26,27]。阻断花色素苷途径上游基因会更有效率,CHS突变也许是使花色缺失最常见的方法^[28,29]。ANS和DFR的突变也可导致花色素缺失^[30,31]。研究表明,通过调节花色素苷生物合成的分支点等方法,也可使花色缺失。将苹果ANR导入烟草可抑制烟草花中CHI和DFR的表达,最终导致花青素含量降低^[32]。本研究中,DFR的低表达和FLS的高表达导致荷兰鸢尾花色变为白色。

4 结论

白色‘玉妃’中,IhDFR1和 IhDFR2的低表达,使花色素苷积累受阻,而与DFR竞争相同底物的IhFLS1在‘玉妃’中高表达,使部分代谢流转向黄酮醇,也在一定程度上限制花色素苷的形成,最终导致花色由蓝紫变为白色。

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