Efficient biosynthesis of glucoraphanin in Brassicaceae crops by genetic engineering

doi:10.12211/2096-8280.2024-031

Abstract

Abstract:

Glucosinolate (GRA), a secondary metabolite of plants, is a Glucosinolate (GSL) derived from methionine. It is relatively stable in nature, and both GRA and its degradation product Sulforaphane (SFN) plays important roles in cancer resistance, neuroprotection, and other broad biological functions and health-benefits, especially, SFN has been widely reported as the best natural product for cancer prevention. GRA has been widely concerned in food nutrition and scientific research. In this paper, we will review the physicochemical properties, sources, biological functions, synthetic pathways, current production status of GRA, and further discuss the potential strategy for efficient biological synthesis of GRA in the future. The synthesis pathway of GRA is complex, involving three stages: side chain elongation, core structure information, and side chain modification. GRA can be converted into SFN and other active compounds by plant myrosinase (MYR) and intestinal microorganisms. Brassicaceae crops such as broccoli have high levels of GRA, and are currently the main source of GRA. However, the cultivation cycle of GRA-rich plant natural resources is long, the yield is unstable, and the extraction rate is low and the development of economical and renewable new resources of GRA will greatly advance its further development and application. With the clarification of the biosynthesis and regulation pathways of GRA, genetic engineering-assisted efficient biological synthesis of GRA shows great potential, suggesting that the possibility of breaking through the mainstream single gene regulatory strategy and aggregating multiple genes and multiple dimensions to enhancement of GRA synthesis. This paper focuses on the goal of the genetic engineering-assisted efficient biosynthesis of GRA in Brassicaceae corps, systematically outlining potential engineering genes at each stage of GRA synthesis and pointing out high-value base crop species from the perspective of enrichment organs, aiming to provide certain ideas and strategies for the future regulation of GRA biosynthesis in plants through transgenic technology and molecular breeding to realize large-scale sustainable production of GRA.

Key words: glucoraphanin, sulforaphane, cancer, genetic engineering, Brassicaceae corps

摘要：

植物次级代谢物萝卜硫苷（Glucoraphanin，GRA）是一种由蛋氨酸衍生的硫代葡萄糖苷（Glucosinolate，GSL），性质相对稳定，其本身及水解后活性产物萝卜硫素（Sulforaphane，SFN）在抵抗癌症、神经保护等方面发挥重要作用，在食品营养和科学研究中受到广泛关注。在本文中，我们将综述GRA的理化性质、来源、生物学功能、合成途径以及当前生产现状，并进一步探讨未来GRA高效生物合成的潜力策略。GRA合成路径复杂，包括侧链延伸、核心结构形成以及侧链修饰三个阶段，可经植物内源黑芥子酶（Myrosinase， MYR）或肠道微生物转化为具有生物活性的SFN等物质。西兰花等十字花科作物中GRA含量较高，是当前GRA的主要来源作物，但其存在种植周期较长、产量不稳，提取率低等问题，开发经济且可再生的GRA新资源将极大的推进GRA开发应用。随着GRA生物合成及调控路径的明晰，基因工程辅助GRA的高效生物合成展现出巨大的潜力，也提示突破主流的单基因调控策略，聚合多基因多维度协同提高GRA合成的潜力。本文聚焦基因工程辅助十字花科作物高效生产GRA这一目标，系统的梳理了GRA合成各阶段的潜在候选基因并从富集部位角度指出了具高应用价值的底盘作物，以期为将来通过基因工程和分子育种技术调控植物中GRA的生物合成，实现GRA大规模可持续生产提供一定的思路和策略。

关键词: 萝卜硫苷（GRA）, 萝卜硫素（SFN）, 癌症, 基因工程, 十字花科作物

CLC Number:

Q812

Xiaoyue LIU, Pandi WANG, Gang Wu, Fang LIU. Efficient biosynthesis of glucoraphanin in Brassicaceae crops by genetic engineering[J]. Synthetic Biology Journal, DOI: 10.12211/2096-8280.2024-031.

刘晓悦, 王盼娣, 吴刚, 刘芳. 基因工程辅助萝卜硫苷在十字花科作物中的高效生物合成[J]. 合成生物学, DOI: 10.12211/2096-8280.2024-031.

Figures/Tables 5

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109	ENDO R, CHIKANO H, ITABASHI E, et al. Large insertion in radish GRS1 enhances glucoraphanin content in intergeneric hybrids, Raphanobrassica (Raphanus sativus L. x Brassica oleracea var. acephala) [J]. Frontiers in Plant Science, 2023, 14.
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119	HöLZL G, REZAEVA B R, KUMLEHN J, et al. Ablation of glucosinolate accumulation in the oil crop Camelina sativa by targeted mutagenesis of genes encoding the transporters GTR1 and GTR2 and regulators of biosynthesis MYB28 and MYB29 [J]. Plant Biotechnology Journal, 2022, 21(1): 189-201.
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124	BELL L, WAGSTAFF C. Glucosinolates, Myrosinase Hydrolysis Products, and Flavonols Found in Rocket (Eruca sativa and Diplotaxis tenuifolia) [J]. Journal of Agricultural and Food Chemistry, 2014, 62(20): 4481-92.
125	HARUN S, M-R ABDULLAH-ZAWAWI, H-H GOH, et al. A Comprehensive Gene Inventory for Glucosinolate Biosynthetic Pathway in Arabidopsis thaliana [J]. Journal of Agricultural and Food Chemistry, 2020, 68(28): 7281-97.
126	SøNDERBY I E, GEU-FLORES F, HALKIER B A. Biosynthesis of glucosinolates – gene discovery and beyond [J]. Trends in Plant Science, 2010, 15(5): 283-90.
127	LIU S, HUANG H, YI X, et al. Dissection of genetic architecture for glucosinolate accumulations in leaves and seeds of Brassica napus by genome‐wide association study [J]. Plant Biotechnology Journal, 2019, 18(6): 1472-84.
128	TANG Y, ZHANG G, JIANG X, et al. Genome-Wide Association Study of Glucosinolate Metabolites (mGWAS) in Brassica napus L [J]. Plants, 2023, 12(3).
129	WEI D, CUI Y, MEI J, et al. Genome‐wide identification of loci affecting seed glucosinolate contents in Brassica napus L [J]. Journal of Integrative Plant Biology, 2018, 61(5): 611-23.
130	BELL L, WAGSTAFF C. Enhancement Of Glucosinolate and Isothiocyanate Profiles in Brassicaceae Crops: Addressing Challenges in Breeding for Cultivation, Storage, and Consumer-Related Traits [J]. Journal of Agricultural and Food Chemistry, 2017, 65(43): 9379-403.
131	BELL L, ORUNA-CONCHA M J, DE HARO-BAILON A. Editorial: Nutritional quality and nutraceutical properties of Brassicaceae (Cruciferae) [J]. Frontiers in Nutrition, 2023, 10.
132	SHEN J, LIU Y, WANG X, et al. A Comprehensive Review of Health-Benefiting Components in Rapeseed Oil [J]. Nutrients, 2023, 15(4).
133	YI G-E, ROBIN A, YANG K, et al. Identification and Expression Analysis of Glucosinolate Biosynthetic Genes and Estimation of Glucosinolate Contents in Edible Organs of Brassica oleracea Subspecies [J]. Molecules, 2015, 20(7): 13089-111.
134	CROCOLL C, MIRZA N, REICHELT M, et al. Optimization of Engineered Production of the Glucoraphanin Precursor Dihomomethionine in Nicotiana benthamiana [J]. Frontiers in Bioengineering and Biotechnology, 2016, 4.
135	ZANG Y-X, KIM J-H, Y-D PARK, et al. Metabolic engineering of aliphatic glucosinolates in Chinese cabbage plants expressing Arabidopsis MAM1, CYP79F1, and CYP83A1 [J]. BMB Reports, 2008, 41(6): 472-8.
136	CHEN S, GLAWISCHNIG E, JøRGENSEN K, et al. CYP79F1 and CYP79F2 have distinct functions in the biosynthesis of aliphatic glucosinolates in Arabidopsis [J]. The Plant Journal, 2003, 33(5): 923-37.
137	REINTANZ B, LEHNEN M, REICHELT M, et al. bus, a Bushy Arabidopsis CYP79F1 Knockout Mutant with Abolished Synthesis of Short-Chain Aliphatic Glucosinolates [J]. The Plant Cell, 2001, 13(2): 351-67.
138	NAUR P, PETERSEN B L, MIKKELSEN M D, et al. CYP83A1 and CYP83B1, Two Nonredundant Cytochrome P450 Enzymes Metabolizing Oximes in the Biosynthesis of Glucosinolates in Arabidopsis [J]. Plant Physiology, 2003, 133(1): 63-72.
139	LIM Y P, ZHANG Y, HUAI D, et al. Overexpression of Three Glucosinolate Biosynthesis Genes in Brassica napus Identifies Enhanced Resistance to Sclerotinia sclerotiorum and Botrytis cinerea [J]. Plos One, 2015, 10(10).
140	HANSEN B G, KLIEBENSTEIN D J, HALKIER B A. Identification of a flavin‐monooxygenase as the S‐oxygenating enzyme in aliphatic glucosinolate biosynthesis in Arabidopsis [J]. The Plant Journal, 2007, 50(5): 902-10.
141	YIN L, CHEN H, CAO B, et al. Molecular Characterization of MYB28 Involved in Aliphatic Glucosinolate Biosynthesis in Chinese Kale (Brassica oleracea var. alboglabra Bailey) [J]. Frontiers in Plant Science, 2017, 8.
142	ARAKI R, HASUMI A, NISHIZAWA O I, et al. Novel bioresources for studies of Brassica oleracea: identification of a kale MYB transcription factor responsible for glucosinolate production [J]. Plant Biotechnology Journal, 2013, 11(8): 1017-27.
143	WANG L, JIANG H, QIU Y, et al. Biochemical Characterization of a Novel Myrosinase Rmyr from Rahnella inusitata for High-Level Preparation of Sulforaphene and Sulforaphane [J]. Journal of Agricultural and Food Chemistry, 2022, 70(7): 2303-11.
144	YE Q, FANG Y, LI M, et al. Characterization of a Novel Myrosinase with High Activity from Marine Bacterium Shewanella baltica Myr-37 [J]. International Journal of Molecular Sciences, 2022, 23(19).
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疾病	摄入方式	机制	效应	参考文献
癌症
脑癌	饲喂西兰花芽提取物	调节Keap1/Nrf2/ARE信号通路，激活细胞抗氧化防御过程	抑制肿瘤生长	[35]
鼻咽癌	添加纯品SFN	抑制 EBV早期裂解蛋白Rta表达，阻断EBV裂解周期	阻断EBV在激活	[13]
鼻咽癌	添加纯品SFN	上调肿瘤抑制因子miRNA-124-3p表达靶向抑制STAT3信号通路表达和磷酸化	抑制癌增殖和转移	[36] [37]
肝癌	添加纯品SFN	诱导 NRF2，重新连接中枢代谢调节调节氨基酸代谢支持谷胱甘肽产生，维持葡萄糖稳态	抗氧化	[38]
肺癌	添加纯品SFN	解聚微管、抑制α-微管蛋白与脂肪酸合酶、乙酰CoA羧化酶、柠檬酸裂解酶相互作用	抑制微管介导的线粒体自噬引起细胞凋亡	[39]
		降低细胞内脂肪酸以及线粒体磷酸含量	抑制癌细胞增值及肿瘤干细胞自我更新	[40]
		调节调节Sonic Hedgehog信号通路和PHC3, 组蛋白修饰降低 miR-616-5p水平	抑制95D和H1299非小细胞肺癌细胞转移	[41]
胃癌	添加纯品SFN	上调Bax/Bcl2蛋白以及细胞色素C、PARP-1等信号蛋白表达,促进丝裂原蛋白激酶（MAPK）JNK和 P-38的磷酸化	促进癌细胞凋亡	[42]
胃癌	添加纯品SFN	下调EGFR（上皮生长因子受体），p-ERK1/2表达	抑制癌细胞转移
胰腺癌	添加纯品SFN	抑制 PI3K/AKT 和 MEK/ERK 通路，激活转录因子 FOXO	诱导细胞周期停滞	[43]
胰腺癌	添加纯品SFN	诱导产生过量活性氧ROS，激活Nrf2-AMPK信号传导途径	抑制癌细胞生长	[44]
结肠癌	添加纯品SFN	靶向降低癌细胞HDAC3活性	表观修饰	[45]
		调节免疫细胞产生的TNFa、IL-1b和IL-6等炎症细胞因子	抗炎活性	[46]
		激活AMPK信号通路	抑制癌细胞生长	[14]
宫颈癌	添加纯品SFN	激活LATS2，阻断Rad51/MDC1修复 DNA 损伤	促进癌细胞凋亡	[47]
前列腺癌	添加纯品SFN	组蛋白H3和H4乙酰化，细胞周期停滞于S和G2/M期	表观修饰，抑制细胞周期	[48]
神经保护
帕金森	添加纯品SFN	Nrf2蛋白、Nrf2mRNA和总谷胱甘肽水平的增加以及神经元组织凋亡的抑制	Nrf2机制调节神经元与小胶质细胞	[49]
阿尔兹海默病	添加纯品SFN	激活Nrf2抗氧化反应元件（ARE），上调细胞对氧化应激的防御，减少神经元丢失	抗氧化	[27, 50]
阿尔兹海默病	添加纯品SFN	促进小胶质细胞从促炎的M1表型向抗炎的M2表型分化，减少神经炎症	抗炎活性	[51]
自闭症	摄入SFN	Nrf2介导的Trx1/TrxR1系统的诱导逆转中性粒细胞损伤	调控细胞周期	[52]
脑内出血	饲喂SFN	SFN激活Nrf2ARE信号通路，发挥抗氧化和抗炎作用，改善脑出血后的神经功能障碍	抗氧化、抗炎	[53]
胎儿神经保护	食用西兰花芽	SFN与酚类物质协调作用，清除自由基及金属络合作用	抗氧化	[28]
其他健康益处
心肌病	饮用SFN水溶液	通过PI3k/Akt/Nrf2信号通路去除砷代谢产生的过量自由基	抗氧化	[32]
心肌病	饮用SFN水溶液	防止砷引起的心脏损伤、氧化应激、线粒体复合物功能障碍	抗氧化
骨质疏松	膳食ITC	诱导NAD（P）H:醌氧化还原酶1的活性，抑制基质中金属蛋白酶1的产生	抗氧化	[33]
肥胖及并发症	进食西兰花等蔬菜	激活Nrf2或有效调节 AMP 激活蛋白激酶	抗氧化、抗炎	[29]
肥胖及并发症	皮下注射SFN	减少机体氧化应激以及炎症生物标志物，促进脂肪组织中的巨噬细胞极化为 M2 表型	调控脂肪族纤维化相关的基因表达	[31]
牛皮癣	腹腔注射SFN	激活 KEAP1-NRF2 通路和减弱炎症信号传导	强抗氧化	[34]

疾病	摄入方式	机制	效应	参考文献
癌症
脑癌	饲喂西兰花芽提取物	调节Keap1/Nrf2/ARE信号通路，激活细胞抗氧化防御过程	抑制肿瘤生长	[35]
鼻咽癌	添加纯品SFN	抑制 EBV早期裂解蛋白Rta表达，阻断EBV裂解周期	阻断EBV在激活	[13]
鼻咽癌	添加纯品SFN	上调肿瘤抑制因子miRNA-124-3p表达靶向抑制STAT3信号通路表达和磷酸化	抑制癌增殖和转移	[36] [37]
肝癌	添加纯品SFN	诱导 NRF2，重新连接中枢代谢调节调节氨基酸代谢支持谷胱甘肽产生，维持葡萄糖稳态	抗氧化	[38]
肺癌	添加纯品SFN	解聚微管、抑制α-微管蛋白与脂肪酸合酶、乙酰CoA羧化酶、柠檬酸裂解酶相互作用	抑制微管介导的线粒体自噬引起细胞凋亡	[39]
		降低细胞内脂肪酸以及线粒体磷酸含量	抑制癌细胞增值及肿瘤干细胞自我更新	[40]
		调节调节Sonic Hedgehog信号通路和PHC3, 组蛋白修饰降低 miR-616-5p水平	抑制95D和H1299非小细胞肺癌细胞转移	[41]
胃癌	添加纯品SFN	上调Bax/Bcl2蛋白以及细胞色素C、PARP-1等信号蛋白表达,促进丝裂原蛋白激酶（MAPK）JNK和 P-38的磷酸化	促进癌细胞凋亡	[42]
胃癌	添加纯品SFN	下调EGFR（上皮生长因子受体），p-ERK1/2表达	抑制癌细胞转移
胰腺癌	添加纯品SFN	抑制 PI3K/AKT 和 MEK/ERK 通路，激活转录因子 FOXO	诱导细胞周期停滞	[43]
胰腺癌	添加纯品SFN	诱导产生过量活性氧ROS，激活Nrf2-AMPK信号传导途径	抑制癌细胞生长	[44]
结肠癌	添加纯品SFN	靶向降低癌细胞HDAC3活性	表观修饰	[45]
		调节免疫细胞产生的TNFa、IL-1b和IL-6等炎症细胞因子	抗炎活性	[46]
		激活AMPK信号通路	抑制癌细胞生长	[14]
宫颈癌	添加纯品SFN	激活LATS2，阻断Rad51/MDC1修复 DNA 损伤	促进癌细胞凋亡	[47]
前列腺癌	添加纯品SFN	组蛋白H3和H4乙酰化，细胞周期停滞于S和G2/M期	表观修饰，抑制细胞周期	[48]
神经保护
帕金森	添加纯品SFN	Nrf2蛋白、Nrf2mRNA和总谷胱甘肽水平的增加以及神经元组织凋亡的抑制	Nrf2机制调节神经元与小胶质细胞	[49]
阿尔兹海默病	添加纯品SFN	激活Nrf2抗氧化反应元件（ARE），上调细胞对氧化应激的防御，减少神经元丢失	抗氧化	[27, 50]
阿尔兹海默病	添加纯品SFN	促进小胶质细胞从促炎的M1表型向抗炎的M2表型分化，减少神经炎症	抗炎活性	[51]
自闭症	摄入SFN	Nrf2介导的Trx1/TrxR1系统的诱导逆转中性粒细胞损伤	调控细胞周期	[52]
脑内出血	饲喂SFN	SFN激活Nrf2ARE信号通路，发挥抗氧化和抗炎作用，改善脑出血后的神经功能障碍	抗氧化、抗炎	[53]
胎儿神经保护	食用西兰花芽	SFN与酚类物质协调作用，清除自由基及金属络合作用	抗氧化	[28]
其他健康益处
心肌病	饮用SFN水溶液	通过PI3k/Akt/Nrf2信号通路去除砷代谢产生的过量自由基	抗氧化	[32]
心肌病	饮用SFN水溶液	防止砷引起的心脏损伤、氧化应激、线粒体复合物功能障碍	抗氧化
骨质疏松	膳食ITC	诱导NAD（P）H:醌氧化还原酶1的活性，抑制基质中金属蛋白酶1的产生	抗氧化	[33]
肥胖及并发症	进食西兰花等蔬菜	激活Nrf2或有效调节 AMP 激活蛋白激酶	抗氧化、抗炎	[29]
肥胖及并发症	皮下注射SFN	减少机体氧化应激以及炎症生物标志物，促进脂肪组织中的巨噬细胞极化为 M2 表型	调控脂肪族纤维化相关的基因表达	[31]
牛皮癣	腹腔注射SFN	激活 KEAP1-NRF2 通路和减弱炎症信号传导	强抗氧化	[34]

俗名	拉丁名	种子	苗（种子刚发芽）	叶	芽	根	参考文献
西兰花	Brassica oleracea var. italica	+/+++	++/+++	++/++	+/+	+/+	[96, 97]
萝卜	Raphanus sativus	+/-	+/-	-/+	+/+	+/-	[96, 98]
白萝卜	xBrassicoraphanus	+/+++	-/++	+/+	+/++	+/++	[96, 98]
甘蓝	Brassica oleracea var. capitata	++/+++	++/+++	++/+	++/++	+/++	[96, 99]
花椰菜	Brassica oleracea var. botrytis	+/+	+/+	N/N	+/+	+/+	[96]
大白菜	Brassica rapa ssp. pekinensis	+/+++	-/+++	+/+	+/+	+/+	[96, 98]
羽衣甘蓝	Brassica oleracea var. acephala	+/++	+/++	++/+	+/++	+/+	[96, 100]
芥菜	Brassica juncea	-/-	+/++	-/N	-/+	-/+	[96, 101]
小白菜	Brassica rapa ssp. Chinensis	+/+	+/+	N/N	+/+	+/++	[96]
欧洲油菜	Brassica napus	+/++	+/+++	+/++	N/N	N/N	[11, 102-104]

俗名	拉丁名	种子	苗（种子刚发芽）	叶	芽	根	参考文献
西兰花	Brassica oleracea var. italica	+/+++	++/+++	++/++	+/+	+/+	[96, 97]
萝卜	Raphanus sativus	+/-	+/-	-/+	+/+	+/-	[96, 98]
白萝卜	xBrassicoraphanus	+/+++	-/++	+/+	+/++	+/++	[96, 98]
甘蓝	Brassica oleracea var. capitata	++/+++	++/+++	++/+	++/++	+/++	[96, 99]
花椰菜	Brassica oleracea var. botrytis	+/+	+/+	N/N	+/+	+/+	[96]
大白菜	Brassica rapa ssp. pekinensis	+/+++	-/+++	+/+	+/+	+/+	[96, 98]
羽衣甘蓝	Brassica oleracea var. acephala	+/++	+/++	++/+	+/++	+/+	[96, 100]
芥菜	Brassica juncea	-/-	+/++	-/N	-/+	-/+	[96, 101]
小白菜	Brassica rapa ssp. Chinensis	+/+	+/+	N/N	+/+	+/++	[96]
欧洲油菜	Brassica napus	+/++	+/+++	+/++	N/N	N/N	[11, 102-104]

	涉及基因	来源物种	受体物种	器官	结果（单位：µmol/g）	参考文献
传统育种
杂交育种	MYB28	Brassica villosa×GD33 X	Beneforte^®	花茎花蕾	GRA 28 DW（2.5-3倍）	[94]
杂交育种	GRS1/grs1	Brassica oleracea var. acephala ×Raphanus sativus L.	Raphanobrassica grs1	叶	GRA 34.1 DW（2倍）	[109]
微生物代谢工程
同工酶替代	BCAT3	Brassica rapa	Escherichia coli BL21（DE3）	菌液	GRA 2-3μg/L	[110, 111]
	GSL-ELONG	Brassica oleracea
	IPMI（LSU1和SSU3）	Arabidopsis thaliana
	IPMDH1	Arabidopsis thaliana
整合外源基因到染色体	CYP79F1	Brassica oleracea	Escherichia coli MG1655		GRA 0.675μg/L	[112]
	CYP83A1	Brassica rapa
	EGT2	Neurospora crassa
	UGT74B1	Arabidopsis thaliana
	ST5c	Brassica rapa
	FMO_GS-OX1	Arabidopsis thaliana
植物代谢工程
过表达	MAM1	Brassica oleracea var. oleracea	Brassica oleracea var. oleracea	茎叶混合物	SFN 增加1.7-3.4倍 FW	[5]
分子标记辅助回交育种	braop2.2/braop2.3	Brassica rapa “R-O-18”（ssp. trilocularis）	Brassica rapa“L58”（ssp. parachinensis）	叶	GRA增加18倍 DW	[113]
过表达AOP1	FMO_GS-OX2	Brassica oleracea var. oleracea	Brassica oleracea var. oleracea	茎叶混合物	SFN增加1.6-2.7倍 FW	[5]
抑制GRA代谢基因	GSL-ALK 基因家族	Brassica napus	Brassica napus	种子	GRA 42.6	[114]
	AOP2	Brassica oleracea var. alboglabra Bailey	Brassica oleracea var. alboglabra Gailan-04	茎	GRA 3.03 DW （3.09倍）	[115]
	AOP2（GSL-ALK）	Brassica juncea	Brassica juncea	种子	GRA 24.1 DW	[116]
	BoaAOP2s	Brassica oleracea var. alboglabra	Brassica oleracea var. alboglabra	叶	GRA 0.082-0.289 FW（11.71-41.29倍）	[117]
过表达转录因子	BoMYB29	Brassica oleracea Winspit	Brassica oleracea DH AG1012	叶	GRA 2.542 FW	[118]
	csmyb28，csmyb29	Camelina sativa	Camelina sativa	种子、根	GSL完全消失	[119]
增加MYR	Myrosianse gene	Brassica oleracea var. oleracea	Brassica oleracea var. oleracea	茎叶混合物	SFN 增加3.7倍 FW	[5]
敲除转运蛋白	gtr2	Arabidopsis thaliana	Arabidopsis thaliana	种子	GSL下降0.48±0.11倍	[79]
	BnaA06.GTR2	Brassica napus	Brassica napus	种子	GSLS下降0.76± 0.0076 倍	[120]
	csgtr1 csgtr2	Camelina sativa	Camelina sativa	种子、根	GSLS减少，种子中减少 0.85-0.88倍	[119]
	gtr1 gtr2	Arabidopsis thaliana	Arabidopsis thaliana	种子、根	GSL种子中几乎消失，根中显著积累	[79]
	Myrosinase-FMO_GS-OX2-MAM1（M-F-A）	Brassica oleracea var. oleracea	Brassica oleracea var. oleracea	茎叶混合物	SFN增加1.8-5.5倍（FW）	[5]
优化基因组合	BCAT3	Arabidopsis thaliana	Nicotiana benthamiana	叶	2.05±0.32 DW（4.74倍）	[121]
	dCGS
	IPMI2
	Aconitase
	CGBP