基于转录组测序的铁皮石斛黄酮代谢途径及相关基因解析

林江波; 王伟英; 邹晖; 戴艺民

doi:10.19303/j.issn.1008-0384.2019.09.004

基于转录组测序的铁皮石斛黄酮代谢途径及相关基因解析

doi: 10.19303/j.issn.1008-0384.2019.09.004

福建省农业科学院亚热带农业研究所，福建　漳州　363005

基金项目: 福建省科技计划公益类专项（2018R1024-3）

详细信息

作者简介:
林江波（1976−），男，硕士，副研究员，主要从事农业生物技术研究（E-mail：345953257@qq.com）

通讯作者:
戴艺民（1969−），男，博士，研究员，主要从事农业生物技术研究（E-mail：dymttcn@163.com）

中图分类号: S 567.239
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出版历程
- 收稿日期: 2019-07-17
- 修回日期: 2019-08-05
- 刊出日期: 2019-09-01

Transcriptome Analysis on Pathway of and Genes Related to Flavonoid Synthesis in Dendrobium officinale

Subtropical Agriculture Research Institute , Fujian Academy of Agricultural Sciences, Zhangzhou, Fujian　363005, China

摘要

摘要: 目的分析铁皮石斛Dendrobium officinale黄酮类化合物的生物合成途径及相关基因，为铁皮石斛黄酮类化合物代谢调控、药用价值的开发研究提供参考。方法利用Illumina HiSeq 4000测序平台对铁皮石斛2个生长阶段茎、叶进行高通量转录组测序，对组装获得的unigenes进行功能注释和黄酮类化合物的生物合成相关基因解析。结果铁皮石斛黄酮类化合物代谢相关Unigenes 48个，涉及14个酶。5个CHS相关Unigenes具有CHS-like保守结构域，活性位点氨基酸残基（Cys-His-Asn）高度保守，丙二酰辅酶A结合位点和产物结合位点的部分氨基酸残基发生变异。CHI(Unigene0013781)属于类型I查尔酮异构酶，异构化6′-羟基查尔酮为5-羟基黄烷酮。表达分析表明，2个生长时期的茎和叶，CHS（Unigene0008250）、CHI（Unigene0013781）和F3H的表达量都高于CHS（Unigene0012884）和C3′H。结论通过对转录组数据分析，共获得铁皮石斛黄酮类化合物代谢相关Unigenes48个，涉及14个酶；5个CHS相关Unigenes中，2个编码查尔酮合酶，3个编码联苄合酶；CHI(Unigene0013781)属于类型I查尔酮异构酶。
- 铁皮石斛 /
- 转录组 /
- 黄酮 /
- 功能注释 /
- 表达 /
- 代谢
Abstract: Objective Pathway and genes related to the flavonoid synthesis in Dendrobium officinale were analyzed for studying on the metabolic regulation and medicinal applications of flavonoids. Method Transcriptomes of the stems and leaves of D. officinale plants at two growth stages were determined by means of Illumina HiSeq 4000 high-throughput sequencing. The assembled unigenes were used for the functional annotation and genetic analysis of the flavonoid biosynthetic pathway. Result There were 48 unigenes involving 14 enzymes found to be associated with the flavonoid metabolism. Among them, the 5 CHS-related unigenes showed a CHS-like conserved domain, highly conserved amino acid residues (Cys-His-Asn) at the active site, and mutated amino acid residues at the malonyl-CoA and product binding sites; and, the CHI (Unigene0013781) was identified as a Type I chalcone isomerase, isomerizing 6'-hydroxychalone to 5-hydroxyflavanone. The expressions of CHS (Unigene0008250), CHI (Unigene0013781) and F3H were higher than those of CHS (Unigene 0012884) and C3′H in the stems and leaves of the plants at both growth stages. Conclusion Forty-eight unigenes involving 14 enzymes associated with the flavonoid metabolism in D. officinale plants were identified by the transcriptome analysis. Of the 5 CHS-related unigenes, two encoded chalcone synthase, and 3 bibenzyl synthase, while the CHI (Unigene0013781) a Type I chalcone isomerase.
- Dendrobium officinale /
- transcriptome /
- flavonoids /
- function annotation /
- expression /
- metabolism

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图 1 铁皮石斛黄酮类化合物生物合成途径

Figure 1. Flavonoid biosynthesis pathway in D. officinale

下载: 全尺寸图片幻灯片

图 2 4个Unigene氨基酸序列的多重比对

注：黑色方块表示活性位点，用ⅰ-ⅲ表示不同位点；黄色方块表示丙二酰辅酶A结合位点，用1-5表示位点位置；红色方块表示产物结合位点，用a-f表示位点位置。

Figure 2. Multi-alignment on amino acid sequences of 4 unigenes

Note: Black squares are active sites ⅰ to ⅲ; yellow squares, malonyl-CoA binding sites 1 to 5; and, red squares, product binding sites a to f.　　　　　

下载: 全尺寸图片幻灯片

图 3 Unigene0013781和其他植物CHI的分子进化树

Figure 3. Molecular phylogenetic trees of Unigene 0013781 and CHI from other plants

下载: 全尺寸图片幻灯片

图 4 铁皮石斛黄酮类生物合成相关Unigene的表达模式

注：T1：生长期茎；T2：生长期叶；T3：成熟期茎；T4：成熟期叶。

Figure 4. Expressions of unigenes related to flavonoid biosynthesis in D. officinale

Note: T1: in stems at growth stage; T2: in leaves at growth stage; T3: in stems at mature stage; T4: in leaves at mature stage.

下载: 全尺寸图片幻灯片

表 1 铁皮石斛转录组黄酮类化合物合成途径相关基因

Table 1. Flavonoids biosynthesis related genes in transcriptone of Dendrobium officinale

KO 编号 KO Number	酶 Enzyme	简写 Abbreviation	Unigene 数 Unigene Number
K10775	苯丙氨酸解氨酶 Phenylalanine ammonia lyase	PAL	4
K01904	4-香豆酰辅酶A连接酶 4-coumaric coenzyme A ligase	4CL	11
K00475	柚皮素3-双加氧酶 Naringin 3-dioxygenase	F3H	1
K00487	反式肉桂酸单加氧酶 Trans-cinnamate monooxygenase	C4H	2
K00588	咖啡基辅酶A O-甲基转移酶 Coffee-based CoA O-methyltransferase	CCoAOMT	5
K00660	查尔酮合酶 Chalcone synthase	CHS	5
K01859	查尔酮异构酶 Chalcone isomerase	CHI	2
K05277	无色花青素双加氧酶 Colorless anthocyanin dioxygenase	LDOX	1
K05278	黄酮醇合酶 Flavonol synthase	FLS	1
K05280	黄酮类3′-单加氧酶 Flavonoid 3′-monooxygenase	F3′H	6
K09754	香豆酰喹酸(香豆杉酯)3′-单加氧酶 Coumarinyl quinoic acid (coumarin) 3′-monooxygenase	C3′H	1
K13065	莽草酸O-羟基肉桂酰基转移酶 Shikimic acid O-hydroxy cinnamoyl transferas	HCT	7
K13082	二氢黄酮醇/黄烷酮4-还原酶 Dihydroflavonol/flavanone 4-reductase	DFR	1
K13083	黄酮类3′,5′-羟化酶 Flavonoids 3′,5′-hydroxylase	F3′5′H	1

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