基于转录组测序的金钱蒲类黄酮生物合成基因的表达分析

陈钰; 马良; 饶雪娥; 何舒澜

doi:10.19303/j.issn.1008-0384.2023.08.006

基于转录组测序的金钱蒲类黄酮生物合成基因的表达分析

doi: 10.19303/j.issn.1008-0384.2023.08.006

福建卫生职业技术学院药学院，福建　福州　350101

基金项目: 福建省教育厅中青年教师教育科研项目（JAT191292）；福建省科技特派员专项（2022年）

详细信息

作者简介:
陈钰（1987 —），女，博士，讲师，主要从事中药鉴定学和中药药理学研究，E-mail：181967453@qq.com

中图分类号: S682
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出版历程
- 收稿日期: 2023-03-06
- 修回日期: 2023-05-15
- 网络出版日期: 2023-09-19
- 刊出日期: 2023-08-28

Expressions of Flavonoid Biosynthesis Genes in Acorus gramineus Determined by Transcriptome Sequencing

Department of Pharmacy, Fujian Health College, Fuzhou, Fujian　350101, China

摘要

摘要: 目的高通量测序获取金钱蒲（Acorus gramineus）7个不同组织转录组信息，为不同组织中类黄酮化学成分合成差异提供分子信息，进而在分子水平上研究金钱蒲不同组织内类黄酮化学成分合成差异。方法利用高通量测序技术平台完成金钱蒲7个不同部位的转录组测序，对unigenes进行功能注释，借助注释结果挖掘类黄酮生物合成通路，筛选通路中的差异表达基因（Differentially expressed genes，DEGs）并进行表达量分析。结果共获得高质量数据39.91～42.97 M，总碱基量为5.98～6.45 Gb，Q30碱基百分比大于94.05%，GC含量为47.81%～50.32%。18616条unigenes在GO数据库中获得62 506个注释，根据功能划分为细胞组分、分子功能及生物过程三大类，分别对应 6、14、21个亚类，大量基因分布在细胞解剖实体、连接、催化活性和细胞过程等亚类中。10124条unigenes 富集在 KEGG 数据库的5大类19个亚类中，从其中筛选出4条类黄酮生物合成途径，14个关键酶，63个差异表达酶基因。这些基因在金钱蒲7个组织中具有表达差异性，表明这些结构基因在类黄酮生物合成过程中于不同部位发挥作用。结论研究结果丰富了金钱蒲的遗传信息，也为进一步解析金钱蒲类黄酮生物合成基因功能提供参考依据。
- 金钱蒲 /
- 转录组 /
- 类黄酮 /
- 基因表达量
Abstract: Objective Transcriptomes of Acorus gramineus tissues were applied to molecularly differentiating flavonoid synthesis genes in the organs. Methods The high-throughput sequencing technology was employed to obtain the transcriptomes of the genes in 7 organs of A, gramineus. Thereby, the functions of unigenes were annotated, the flavonoid biosynthesis pathway deciphered, the differentially expressed genes (DEGs) in the pathway screened, and an expression analysis performed. Results A total of 39.91 to 42.97 M of high-quality data were secured with 5.98–6.45 Gb bases, more than 94.05% Q30 bases, and a GC content of 47.81%–50.32%. From the GO database, 62506 were annotated to classify the 18616 unigenes into 3 functional groups that included cellular components, molecular functions, and biological processes corresponding to 6, 14, and 21 subcategories, respectively. Numerous genes were distributed in the subcategories, such as cellular anatomical entities, connections, catalytic activities, and cellular processes. There were 10124 unigenes enriched in the 5 major categories and 19 subclasses of the KEGG database. From which, 4 flavonoid biosynthesis pathways, 14 key enzymes, and 63 DEGs were screened. The transcriptomes in 7 tissues of A. gramineus obtained in this study led to the identification of 63 DEGs involved in 4 flavonoid biosynthesis pathways. These genes differed in expressions in the organs indicating their diversified roles in the flavonoid biosynthesis in A. gramineus Conclusion The research results have enriched the genetic information of A. gramineus and provided a reference for further elucidating the functional genes involved in the biosynthesis of flavonoids in A. gramineus.
- Acorus gramineus /
- transcriptome /
- flavonoids /
- expression analysis

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图 1 金钱蒲7个不同组织

A：叶；B：花；C：花序；D：果序；E：果实；F：根；G茎。

Figure 1. Seven sampled organs of A. gramineus

A: leaves; B: flower; C: inflorescence; D: infructescence; E: fruit; F: root; G: stem.

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图 2 金钱蒲转录组unigenes的GO功能分类

Figure 2. GO function classification on unigenes of A. gramineus transcriptomes

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图 3 金钱蒲转录组基因的KEGG代谢通路分类

Figure 3. KEGG metabolic pathways of unigenes of A. gramineus transcriptomes

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图 4 金钱蒲类黄酮生物合成差异表达基因热图

A：苯丙素合成中的差异表达基因；B：类黄酮化合物生物合成中的差异表达基因；C：异黄酮代谢通路中的差异表达基因；D：黄酮和黄酮醇生物合成中的差异表达基因。 I：花序；II：花；III：叶；IV：茎；V：根；VI：果序；VII：果。高表达水平为红色，低表达水平为蓝色。

Figure 4. Heat map of DEGs in flavonoid biosynthesis of A. gramineus

A: DEGs in phenylpropanoid synthesis; B: DEGs in flavonoid synthesis; C: DEGs in isoflavone metabolic pathway；D: DEGs in biosynthesis of flavonoids and flavonols. I: inflorescence; II: flowers; III: leaves; IV: stems; V: roots; VI: infructescence; VII: fruits. High levels shown in red, and low in blue.

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表 1 转录组过滤数据统计和参考基因组比对结果

Table 1. Statistically filtered transcriptome data vs. reference genomes

样品名称 Sample name	Clean reads 数量 Number of clean reads/M	高质量总碱基量 Total clean bases/Gb	Q30碱基百分比 Clean reads Q30/%	GC含量 GC content%	比对上参考基因组的 Clean reads 比例 Total mapping/%	比对上参考基因组唯一位置的 Clean reads 比例 Uniquely mapping/%
花序 Inflorescence	42.74	6.41	93.68	49.55	86.50	50.81
花 Flower	42.76	6.41	93.56	50.32	86.26	49.96
叶 Leaf	42.69	6.40	94.05	49.67	82.74	47.88
茎 Stem	42.77	6.41	93.87	50.07	85.80	51.13
根 Root	42.97	6.45	93.55	47.81	87.49	50.39
果序 Infructescence	42.86	6.43	93.99	48.73	77.57	43.82
果 Fruit	39.91	5.98	93.31	50.30	83.38	46.93

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表 2 金钱蒲转录组类黄酮生物合成途径中的酶基因

Table 2. Genes of enzymes inA. gramineus transcriptomes involved flavonoid synthesis pathways

代谢通路 Metabolic pathway	ko编号 Ko ID	酶 Enzyme	缩写 Abbreviation	差异基因数量 Number of DEGs
苯丙素合成 Phenylpropanoid biosynthesis	ko00940	苯丙氨酸解氨酶 Phenylalanine ammonia-lyase	PAL	6
		咖啡酰辅酶A-O-甲基转移酶 Trans-cinnamate 4-monooxygenase	CCOMT	4
		4-香豆酸-CoA 连接酶 4-coumarate--CoA ligase	4CL	4
类黄酮化合物的生物合成 Flavonoid biosynthesis	ko00941	查尔酮异构酶 chalcone isomerase	CHI	11
		查尔酮合成酶 Chalcone synthase	CHS	5
		柚皮素3-双加氧酶 Naringenin 3-dioxygenase	F3H	4
		二氢黄酮醇4-还原酶 Dihydroflavonol 4-reductase	DFR	1
		花青素合成酶 Anthocyanidin synthase	ANS	2
异黄酮代谢通路 Isoflavone metabolic pathway	ko00943	2-羟基异黄酮脱水酶 2-hydroxyisoflavanone dehydratase	HIDH	6
		4’-甲氧基异黄酮2’-羟化酶 4'-methoxyisoflavone 2'-hydroxylase	CYP81E	5
		异黄酮7-O-葡萄糖苷-6’’-O-丙二酰基转移酶 isoflavone 7-O-glucoside-6’-O-malonyltransferase	IF7MAT	5
黄酮和黄酮醇的生物合成 Flavone and flavonol biosynthesis	ko00944	黄酮醇3-O-葡萄糖基转移酶 Flavonol 3-O-glucosyltransferase	F3G	1
		黄酮醇-3-O-葡萄糖苷L-鼠李糖基转移酶 Flavonol-3-O-glucoside L-rhamnosyltransferase	F3GL	2
		类黄酮3’-单加氧酶 Flavonoid 3'-monooxygenase	CYP75B1	6

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