花生ω-3脂肪酸脱氢酶基因在拟南芥中异源表达及转录组分析

doi:10.3724/SP.J.1006.2025.55005

作物学报 ›› 2025, Vol. 51 ›› Issue (12): 3251-3265.doi: 10.3724/SP.J.1006.2025.55005

• 作物遗传育种·种质资源·分子遗传学 • 上一篇下一篇

花生ω-3脂肪酸脱氢酶基因在拟南芥中异源表达及转录组分析

许静¹(), 毕竞男¹, 殷祥贞¹, 赵健鑫¹^,², 赵旭红¹, 潘丽娟¹, 陈娜¹, 姜骁¹, 马俊卿¹, 殷冬梅³, 迟晓元¹^,^*()

¹山东省花生研究所, 山东青岛 266100
²青岛科技大学生物工程学院, 山东青岛 266042
³河南农业大学农学院, 河南郑州 450002

收稿日期:2025-01-08 接受日期:2025-07-09 出版日期:2025-12-12 网络出版日期:2025-07-15
通讯作者: *迟晓元, E-mail: chi000@126.com
作者简介:E-mail: xu_jing_cool@yeah.net
基金资助:
本研究由财政部和农业农村部国家现代农业产业技术体系建设专项(CARS-13);新疆维吾尔自治区重大科技专项(2022A02008-3);山东省农业科学院农业科技创新工程项目(CXGC2025F19);花生所基础研究任务青年基金一般任务项目(CXGC2025C19);泰山学者工程项目(tstp20240523);泰山学者工程项目(tsqn202312292);山东省自然科学基金项目(ZR2023QC177);山东省自然科学基金项目(ZR2023QC146);山东省重点研发计划项目(2024LZGC035);农业农村部油料作物生物学与遗传育种重点实验室项目(KF2024007);青岛市自然科学基金青年项目(25-1-1-38-zyyd-jch)

Heterologous expression of peanut ω-3 fatty acid desaturase genes in Arabidopsis thaliana and transcriptome analysis

XU Jing¹(), BI Jing-Nan¹, YIN Xiang-Zhen¹, ZHAO Jian-Xin¹^,², ZHAO Xu-Hong¹, PAN Li-Juan¹, CHEN Na¹, JIANG Xiao¹, MA Jun-Qing¹, YIN Dong-Mei³, CHI Xiao-Yuan¹^,^*()

¹Shandong Peanut Research Institute, Qingdao 266100, Shandong, China
²College of Biological Engineering, Qingdao University of Science and Technology, Qingdao 266042, Shandong, China
³College of Agriculture, Henan Agricultural University, Zhengzhou 450002, Henan, China

Received:2025-01-08 Accepted:2025-07-09 Published:2025-12-12 Published online:2025-07-15
Contact: *E-mail: chi000@126.com
Supported by:
China Agriculture Research System of MOF and MARA(CARS-13);Major Scientific and Technological Project in Xinjiang(2022A02008-3);Innovation Project of SAAS(CXGC2025F19);Peanut Institute Basic Research Tasks Youth Fund General Tasks(CXGC2025C19);Taishan Scholars Program(tstp20240523);Taishan Scholars Program(tsqn202312292);Natural Science Fund of Shandong Province(ZR2023QC177);Natural Science Fund of Shandong Province(ZR2023QC146);Key R&D Program of Shandong Province, China(2024LZGC035);Open Project of Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs(KF2024007);Qingdao Natural Science Foundation Youth Project(25-1-1-38-zyyd-jch)

摘要/Abstract

摘要： ω-3脂肪酸脱氢酶(ω-3 FAD)催化亚油酸合成α-亚麻酸。亚麻酸为多元不饱和脂肪酸, 对人体的成长及发育、免疫反应等具有保健功能。为了探究ω-3 FAD在花生生长发育中的作用, 本研究从花生中克隆了4个ω-3脂肪酸脱氢酶基因AhFAD3a、AhFAD3c、AhFAD7a和AhFAD7d, 并将它们共转化到拟南芥中。结果表明, 转基因拟南芥异源表达株系种子的脂肪酸组分与野生型存在较大差异, 总脂肪酸含量均显著增加, 增幅为4.50%~9.00%。亚细胞定位分析表明, AhFAD3a和AhFAD3c定位于内质网上, AhFAD7a和AhFAD7d定位于叶绿体中。GO注释和KEGG富集结果显示, 差异表达的基因富集在多种生物学代谢进程, 包括脂质代谢、氨基酸代谢、碳水化合物代谢等, 11个上调表达基因富集在α-亚油酸代谢通路, 共筛选到15个差异表达基因参与脂肪酸代谢途径。研究结果为花生脂肪酸代谢的分子机制提供理论基础, 为花生品质的遗传改良提供了新线索。

关键词: ω-3脂肪酸脱氢酶, 拟南芥, 多基因共表达, 转录组学, 脂肪酸代谢

Abstract:

The enzyme omega-3 fatty acid dehydrogenase (ω-3 FAD) catalyzes the conversion of linoleic acid to alpha-linolenic acid, a polyunsaturated fatty acid essential for human growth, development, and immune function. To explore the role of ω-3 FAD in peanut growth and development, four ω-3 fatty acid dehydrogenase genes—AhFAD3a, AhFAD3c, AhFAD7a, and AhFAD7d—were cloned from peanut and co-transformed into Arabidopsis thaliana in this study. The results showed that the fatty acid profiles of seeds from transgenic overexpression lines differed significantly from those of the wild type, with total fatty acid content increased by 4.50% to 9.00%. Subcellular localization analysis revealed that AhFAD3a and AhFAD3c were localized to the endoplasmic reticulum, while AhFAD7a and AhFAD7d were targeted to the chloroplast. GO annotation and KEGG pathway enrichment analysis indicated that the differentially expressed genes were significantly enriched in various biological processes, including lipid, amino acid, and carbohydrate metabolism. Notably, 11 upregulated genes were involved in the α-linolenic acid metabolic pathway, and a total of 15 differentially expressed genes associated with fatty acid metabolism were identified. These findings provide a theoretical foundation for understanding the molecular mechanisms of fatty acid metabolism in peanut and offer new insights for the genetic improvement of peanut quality.

Key words: ω-3 fatty acid dehydrogenase, Arabidopsis thaliana, co-expression of multiple gene, transcriptomics, fatty acid metabolism

许静, 毕竞男, 殷祥贞, 赵健鑫, 赵旭红, 潘丽娟, 陈娜, 姜骁, 马俊卿, 殷冬梅, 迟晓元. 花生ω-3脂肪酸脱氢酶基因在拟南芥中异源表达及转录组分析[J]. 作物学报, 2025, 51(12): 3251-3265.

XU Jing, BI Jing-Nan, YIN Xiang-Zhen, ZHAO Jian-Xin, ZHAO Xu-Hong, PAN Li-Juan, CHEN Na, JIANG Xiao, MA Jun-Qing, YIN Dong-Mei, CHI Xiao-Yuan. Heterologous expression of peanut ω-3 fatty acid desaturase genes in Arabidopsis thaliana and transcriptome analysis[J]. Acta Agronomica Sinica, 2025, 51(12): 3251-3265.

图/表 10

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表1

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表2

脂肪酸代谢通路差异表达基因注释及表达倍数"

基因编号 Gene ID	基因注释 Gene annotation	KEGG通路 KEGG pathway	调控 Regulation	差异表达倍数 Fold change
AT1G17420	脂氧合酶 Lipoxygenase	亚油酸代谢; α-亚麻酸代谢 Linoleic acid metabolism; alpha-linolenic acid metabolism	上调 up	1.51
AT1G17990	FMN氧化还原酶类 FMN oxidoreductase class	α-亚麻酸代谢 Alpha-linolenic acid metabolism	上调 up	1.65
AT1G19640	茉莉酸羧甲基转移酶 Jasmonic acid carboxyl methyltransferase	α-亚麻酸代谢 Alpha-linolenic acid metabolism	上调 up	1.57
AT2G33150	乙酰-CoA酰基转移酶1 Acetyl-CoA acyltransferase 1	脂肪酸降解; 缬氨酸、亮氨酸和异亮氨酸降解; α-亚麻酸代谢; 不饱和脂肪酸的生物合成; 脂肪酸代谢; 过氧化物酶体 Fatty acid degradation; valine, leucine and isoleucine degradation; alpha-linolenic acid metabolism; biosynthesis of unsaturated fatty acids; fatty acid metabolism; peroxisome	上调 up	1.59
AT3G01420	α-双加氧酶1 Alpha-dioxygenase 1	α-亚麻酸代谢 Alpha-linolenic acid metabolism	上调 up	2.29
AT3G03480	乙酰辅酶a:(Z)-3-己烯-1-醇乙酰转移酶 Acetyl CoA:(Z)-3-hexen-1-ol acetyltransferase	α-亚麻酸代谢 Alpha-linolenic acid metabolism	上调 up	1.77
AT3G25780	烯氧化物环化酶 Allene oxide cyclase	α-亚麻酸代谢 Alpha-linolenic acid metabolism	上调 up	1.62
AT4G13550	质体单半乳糖二酰基甘油脂肪酶 Plasmatic monogalactosyldiacylglycerol lipase	甘油磷脂代谢; α-亚麻酸代谢 Glycerophospholipid metabolism; alpha-linolenic acid metabolism	上调 up	1.75
AT4G18550	细胞质酰基水解酶Cytosolic acyl hydrolase	甘油磷脂代谢; α-亚麻酸代谢 Glycerophospholipid metabolism; alpha-linolenic acid metabolism	上调 up	2.38
AT5G04040	三酰基甘油脂肪酶 TAG lipase	类固醇的生物合成; 甘油磷脂代谢; 醚脂代谢; 花生四烯酸代谢; 亚油酸代谢; α-亚麻酸代谢 Steroid biosynthesis; glycerolipid metabolism; glycerophospholipid metabolism; ether lipid metabolism; arachidonic acid metabolism; linoleic acid metabolism; alpha-linolenic acid metabolism	上调 up	1.55
AT5G65110	酰基辅酶A氧化酶 Acyl-CoA oxidase	脂肪酸降解; β-丙氨酸代谢; α-亚麻酸代谢; 丙酸盐代谢; 不饱和脂肪酸的生物合成; 碳代谢; 脂肪酸代谢; 过氧物酶体 Fatty acid degradation; beta-alanine metabolism; alpha-linolenic acid metabolism; propanoate metabolism; biosynthesis of unsaturated fatty acids; carbon metabolism; fatty acid metabolism; peroxisome	上调 up	1.50

表2

表3

脂肪酸代谢相关基因注释及表达倍数"

基因编号 Gene ID	基因注释 Gene annotation	KEGG 通路 KEGG pathway	调控 Regulation	差异表达倍数 Fold change
AT1G02205	醛脱羰基酶 Aldehyde decarbonylase	角质、木栓质和蜡质的生物合成 Cutin, suberine and wax biosynthesis	下调 down	0.65
AT1G07720	3-酮酰-辅酶A合成酶 3-ketoacyl-CoA synthase	脂肪酸延长; 植物病原菌互作 Fatty acid elongation; plant-pathogen interaction	下调 down	0.64
AT1G23800	醛脱氢酶 Aldehyde dehydrogenase	糖酵解/糖异生; 抗坏血酸和醛酸代谢; 脂肪酸降解; 缬氨酸、亮氨酸和异亮氨酸降解; 赖氨酸降解; 精氨酸和脯氨酸代谢; 组氨酸代谢; 色氨酸代谢; β-丙氨酸代谢; 甘油磷脂代谢; 丙酮酸代谢; 泛酸和CoA的生物合成; 柠檬烯和蒎烯降解 Glycolysis/Gluconeogenesi; ascorbate and aldarate metabolism; fatty acid degradation; valine, leucine and isoleucine degradation; lysine degradation; arginine and proline metabolism; histidine metabolism; tryptophan metabolism; beta-alanine metabolism; glycerolipid metabolism; pyruvate metabolism; pantothenate and CoA biosynthesis; limonene and pinene degradation	上调 up	1.76
AT1G43800	植物硬脂酰-酰基-载体蛋白去饱和酶 Plant stearoyl-acyl-carrier-protein desaturase	脂肪酸的生物合成; 不饱和脂肪酸的生物合成; 脂肪酸代谢 Fatty acid biosynthesis; biosynthesis of unsaturated fatty acids; fatty acid metabolism	上调 up	1.65
AT1G75000	GNS1/SUR4膜蛋白家族GNS1/SUR4 Membrane protein family	脂肪酸延长 Fatty acid elongation	上调 up	2.29
AT2G33150	乙酰-CoA酰基转移酶 1 Acetyl-CoA acyltransferase 1	脂肪酸降解; 缬氨酸、亮氨酸和异亮氨酸降解; α-亚麻酸代谢; 不饱和脂肪酸的生物合成; 脂肪酸代谢; 过氧物酶体 Fatty acid degradation; valine, leucine and isoleucine degradation; alpha-linolenic acid metabolism; biosynthesis of unsaturated fatty acids; fatty acid metabolism; peroxisome	上调 up	1.59
AT2G33380	过氧化物酶 Peroxygenase	木质素、亚木质素和蜡的生物合成 Cutin, suberine and wax biosynthesis	上调 up	1.71
AT2G46720	3-酮酰-辅酶A合成酶 3-ketoacyl-CoA synthase	脂肪酸伸长; 植物病原菌互作 Fatty acid elongation; plant-pathogen interaction	下调 down	0.51
AT3G06810	酰-CoA脱氢酶 Acyl-CoA dehydrogenase	脂肪酸降解; 缬氨酸、亮氨酸和异亮氨酸降解; 脂肪酸代谢 Fatty acid degradation; valine, leucine and isoleucine degradation; fatty acid metabolism	上调 up	1.64
AT3G44540	醇形成脂肪酸酰-CoA还原酶 Alcohol-forming fatty acyl-CoA reductase	木质素、亚木质素和蜡的生物合成; 过氧物酶体 Cutin, suberine and wax biosynthesis; peroxisome	上调 up	2.81
AT5G08250	脂肪酸ω-羟化酶 Fatty acid omega-hydroxylase	木质素、亚木质素和蜡的生物合成 Cutin, suberine and wax biosynthesis	上调 up	3.22
AT5G22500	醇形成脂肪酸酰-CoA还原酶 Alcohol-forming fatty acyl-CoA reductase	木质素、亚木质素和蜡的生物合成; 过氧物酶体 Cutin, suberine and wax biosynthesis; peroxisome	上调 up	1.74
AT5G47350	棕榈酰-蛋白硫酯酶 Palmitoyl-protein thioesterase	脂肪酸伸长; 脂肪酸代谢 Fatty acid elongation; fatty acid metabolism	下调 down	0.58
AT5G57800	醛脱羰基酶 Aldehyde decarbonylase	木质素、亚木质素和蜡的生物合成 Cutin, suberine and wax biosynthesis	上调 up	1.63
AT5G65110	酰基-CoA氧化酶 Acyl-CoA oxidase	脂肪酸降解; β-丙氨酸代谢; α-亚麻酸代谢; 丙酸代谢; 不饱和脂肪酸的生物合成; 碳代谢; 脂肪酸代谢; 过氧化物酶体 Fatty acid degradation; beta-alanine metabolism; alpha-linolenic acid metabolism; propanoate metabolism; biosynthesis of unsaturated fatty acids; carbon metabolism; fatty acid metabolism; peroxisome	上调 up	1.50

表3

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样品名称 Sample name	过滤后数据 Clean read	总碱基数 Clean base	GC含量 GC content (%)	Q20占比 Proportion of Q20 (%)	Q30占比 Proportion of Q30 (%)
FAD1	27,801,157	8,304,078,002	46.79	97.45	93.25
FAD2	20,116,880	6,022,025,662	46.78	97.51	93.23
FAD3	30,223,351	9,034,644,418	46.59	97.02	92.35
WT1	25,057,355	7,495,402,486	46.26	97.42	93.18
WT2	26,695,017	7,973,829,106	46.70	96.95	92.43
WT3	25,050,832	7,485,133,406	46.68	97.39	93.15

花生ω-3脂肪酸脱氢酶基因在拟南芥中异源表达及转录组分析

Heterologous expression of peanut ω-3 fatty acid desaturase genes in Arabidopsis thaliana and transcriptome analysis

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