|Table of Contents|

Gene Target Design and Functional Analysis of Soybean DGAT1/2 Genes(PDF)

《大豆科学》[ISSN:1000-9841/CN:23-1227/S]

Issue:
2022年04期
Page:
438-447
Research Field:
Publishing date:

Info

Title:
Gene Target Design and Functional Analysis of Soybean DGAT1/2 Genes
Author(s):
YANG CenCHEN Li-yuLIAO Chun-meiKONG Fan-jiang
(Innovation Center of Molecular Genetics and Evolution, School of Life Sciences, Guangzhou University, Guangzhou 510006, China)
Keywords:
soybean DGAT1 DGAT2 gene expression transient expression in tobacco leaves CRISPR/Cas9 knockout target soybean hair root tansformation
PACS:
-
DOI:
10.11861/j.issn.1000-9841.2022.04.0438
Abstract:
Diacylglyceryl transferase (DGAT) is a speed-limiting enzyme that catalyzes triacylglycerol (TAG) from diacylglycerol (DAG). TAG is the main storage form of plant seed oil. Excessive TAG in human will only not cause hyperlipidemia and hyperglycemia, but also overload metabolism of liver function, resulting in impaired liver function, increased serum transaminase content and eventually formed fatty liver. While, DAG is more beneficial to human health. In order to lay the groundwork of acquire the mutant and understanding the function of DGAT1/2 genes, using quantitative PCR to detect tissue expression patterns of the eight GmDGAT1/2 genes (DGAT1a,DGAT1b,DGAT1c,DGAT2a,DGAT2b,DGAT2c,DGAT2d and DGAT2e), expressed GmDGAT1c and GmDGAT2b separately in tobacco leaves. And we constructed the knockout vectors of DGAT1/2 and transformed them into soybean hair roots for target detection by CRISPR/Cas9 technology. The results showed that GmDGAT1/2 were expressed in all detected tissues of soybeans, and the abundance of expression in seeds was higher than that in other tissues. And the expression of them in the seeds was the highest at seeds of mature stage (R8). The fat content of tobacco leaves, which expressed GmDGAT1c and GmDGAT2b separately was increased by 32.53% and 25.90%. In addition, we obtained vector GmDGAT1-Cas9 containing effective targets DGAT1a and DGAT1c, and vector GmDGAT2-Cas9 containing effective targets DGAT2a, DGAT2b, DGAT2c, DGAT2d and DGAT2e.

References:

[1]NAN H, LU S, FANG C, et al. Molecular breeding of a high oleic acid soybean line by integrating natural variations[J]. Molecular Breeding, 2020, 40(9): 1-10. [2]邱玲. 中国食用植物油贸易竞争力研究[D]. 哈尔滨: 东北农业大学, 2017. (QIU L. Study on trade competitiveness of Chinese edible vegetable oil[D]. Harbin: Northeast Agricultural University, 2017.)[3]任波, 李毅. 大豆种子脂肪酸合成代谢的研究进展[J]. 分子植物育种, 2005(3): 301-306. (REN B, LI Y. Research advances on fatty acid biogynthesis metabolism in soybean seed[J]. Molecular Breeding, 2005(3): 301-306.)[4]〖ZK(〗MURASE T, MIZUNO T, OMACHI T, et al. Dietary diacylglycerol suppresses high fat and high sucrose diet-induced body fat accumulation in C57BL/6J mice[J]. Journal of Lipid Research, 2001, 42(3): 372-378. [5]〖ZK(〗NAGAO〖KG(0.9mm〗 T, WATANABE H, GOTO N, et al. Dietary diacylglycerol suppresses accumulation of body fat compared to triacylglycerol in men in a double-blind controlled trial[J]. The Journal of Nutrition, 2000, 130(4): 792-797. [6]乔睿. 怎样有效降低甘油三酯[N]. 健康报, 2019-06-04(7). (QIAO R. How to reduce triglycerides effectively [N]. Health, 2019-06-04(7).)[7]李熠阳, 王远亮. 甘油二酯的功能及安全性评价研究进展[J]. 食品与机械, 2012, 28(3): 255-257, 261. (LI Y Y, WANG Y L. Functions and safety evaluation of diacylglycerol[J]. Food & Machinery, 2012, 28(3): 255-257, 261.)[8]LI R, HATANAKA T, YU K, et al. Soybean oil biosynthesis: Role of diacylglycerol acyltransferases[J]. Functional & Integrative Genomics, 2013,13: 99-113. [9]仲晓芳, 钱雪艳, 牛陆, 等. 转基因技术对提高大豆油脂和油酸含量的作用[J]. 大豆科技, 2019 (6): 27-29. (ZHONG X F, QIAN X Y, NIU L, et al. Effect of transgenic technology on improving soybean oil and oleic acid content[J]. Soybean Science & Technology, 2019(6): 27-29.)[10]LARDIZABAL K, EFFERTZ R, LEVERING C, et al. Expression of umbelopsis ramanniana DGAT2A in seed increases oil in soybean[J]. Plant Physiology, 2008, 148(1): 89-96. [11]边妙, 郭葳, 周新安, 等. 转基因科普系列-转基因技术提高大豆油脂品质[J]. 大豆科技, 2018(4): 54-58. (BIAN M, GUO W, ZHOU X A, et al. Improveing soybean oil quality by GM technology[J]. Soybean Science and Technology, 2018(4): 54-58.)[12]张飞, 高秀清, 张靖洁, 等. 种子特异表达异源DGAT1基因提高大豆种子含油量和营养品质[J]. 生物工程学报, 2018, 34(9): 1478-1490.(ZHANG F, GAO X Q, ZHANG J J, et al. Seed-specific expression of heterologous gene DGAT1 increase soybean seed oil content and nutritional quality[J]. Chinese Journal of Biotechnology, 2018, 34(9): 1478-1490.)[13]赵翠格, 刘頔, 李凤兰, 等. 植物种子油脂的生物合成及代谢基础研究进展[J]. 种子, 2010, 29(4): 56-62. (ZHAO C G, LIU D, LI F L, et al. Advances in research on seed oil biosynthesis and basal metabolism[J]. Seed, 2010, 29(4): 56-62.)[14]黄卓烈, 朱利泉. 生物化学[M]. 北京:中国农业出版社, 2004: 226-232. (HUANG Z L, ZHU L Q. Biochemistry[M]. Beijing: China Agriculture Press, 2004: 226-232.)[15]马三梅. 植物中从二酰甘有到三酰甘油的两条合成新途径[J]. 生命的化学, 2006, 26(1): 65-66. (MA S M. Two new pathways from diacylglycerol to triacylglycerol in plants[J]. Chemistry of Life, 2006, 26(1): 65-66.)[16]陶芬芳, 邢蔓, 岳宁燕, 等. 植物甘油三酯合成相关基因研究进展[J]. 作物研究, 2017, 31(3): 330-336. (TAO F F, XING M, YUE N Y, et al. Research advances of genes related to plant triacylglycerol synthesis[J]. Crop Research, 2017, 31(3): 330-336.)[17]DAHLQVIST A, STAHL U, LENMAN M, et al. Phospholipid: Dia-cylglycerol acyltransferase: An enzyme that catalyzes the acyl-CoA-independent formation of triacylglycerol in yeast and plants[J]. Proceedings of the National Academy of Sciences of the United States of America, 2000, 97(12): 6487-6492. [18]LU L, WEI W, LI Q T, et al. A transcriptional regulatory module controls lipid accumulation in soybean[J]. New Phytologist, 2021, 231(2): 661-678. [19]郑玲. 花生二酰甘油酰基转移酶(AhDGAT)基因家族的功能与调控研究[D]. 济南: 山东大学, 2018. (ZHENG L. Function and regulation research of peanut Diacylgycerol Acyltransferase(AhDGAT)gene family[D]. Jinan: Shandong University, 2018.)[20]苑丽霞, 毛雪, 高昌勇, 等. 种子特异表达二酰甘油酰基转移酶1基因(VgDGAT1)提高亚麻荠种子油脂积累 [J]. 植物生理学报, 2015, 51 (5): 668-678. (YUAN L X, MAO X, GAO C Y, et al. Seed-specific over-expression of a Diacylglycerol Acyltransferase 1 gene (VgDGAT1) increase seed oil accumulation in Camelina sativa[J]. Plant Physiology Communications, 2015, 51(5): 668-678.)[21]任国鹏, 葛丽萍, 孙超超, 等. 续随子二酰甘油酰基转移酶2基因(ElDGAT2)克隆与功能分析[J]. 植物生理学报, 2019, 55(8): 1156-1166.(REN G P, GE L P, SUN C C, et al. Cloning and functional analysis of acyl-CoA:Diacylglycerol acyltransferase 2 gene (ElDGAT2) in Euphorbia lathyris[J]. Plant Physiology Communications, 2019, 55(8): 1156-1166.)[22]袁秀云, 田云芳, 张燕, 等. 油用牡丹PEPC基因的克隆及表达分析[J]. 中国油料作报, 2019, 12(10): 1-9. (YUAN X Y, TIAN Y F, ZHANG Y, et al. Cloning and expression analysis of PEPC gene from Paeonia ostii[J]. Chinese Journal of Oil Crop Sciences, 2019, 12(10): 1-9.)[23]孟祥河. 功能性甘油二酯的酶促酯化合成及其减肥功能的研究[D]. 无锡: 江南大学, 2004. (MENG X H. Study on enzymatic synthesis of functional lipid and its antiobesity characteristics[D]. Wuxi: Jiangnan University, 2004.)[24]李聪, 曹文广. CRISPR/Cas9介导的基因编辑技术研究进展[J]. 生物工程学报, 2015, 31(11): 1531-1542. (LI C, CAO W G. Advances in CRISPR/Cas9-mediated gene editing[J]. Chinese Journal of Biotechnology, 2015, 31(11): 1531-1542.)[25]MOJICA F, DíEZ-VILLASE O R C, GARCíA-MARTíNEZ J, et al. Intervening sequences of regularly spaced prokaryotic repeats derive from foreign genetic elements[J]. Journal of Molecular Evolution, 2005, 60(2): 174-182.[26]马兴亮, 刘耀光. 植物CRISPR/Cas9基因组编辑系统与突变分析[J]. 遗传, 2016, 38(2): 118-125. (MA X L, LIU Y G. CRISPR/Cas9-based genome editing systems and the analysis of targeted genome mutations in plants[J]. Hereditas, 2016, 38(2): 118-125.)[27]CHEN L, NAN H A, KONG L, et al. Soybean AP1 homologs control flowering time and plant height[J]. Journal of Integrative Plant Biology, 2020, 62(12): 1868-1879.[28]姜丽静,马春玲,车淑静. 大豆育种中转基因技术的研究进展[J]. 现代化农业, 2017(12): 15-16. (JIANG L J, MA C L, CHE S J. Research progress of transgenic technology in soybean breeding[J]. Modernizing Agriculture, 2017(12): 15-16.)[29]李泰, 杜浩, 黎永力, 等. 大豆BBX32基因生物信息学分析及基因编辑靶点设计[J]. 大豆科学, 2021, 40(5): 602-611. (LI T, DU H, LI Y L, et al. Bioinformatics analysis and gene editing target design of BBX32 gene in soybean[J]. Soybean Science, 2021, 40(5): 602-611.)[30]黎永力, 杜浩, 李泰, 等. 大豆FUL基因家族进化规律分析及基因编辑靶点鉴定[J]. 植物遗传资源学报, 2021, 22(4): 1120-1132. (LI Y L, DU H, LI T, et al. Molecular evolution of FUL family genes and identification of gene editing targets in soybean[J]. Journal of Plant Genetic Resources, 2021, 22(4): 1120-1132.)[31]朱红霞, 胡利宗, 邓小莉, 等. 三种豆科植物DGAT1基因家族的分子特征与进化分析[J]. 生物技术通报, 2011(10): 163-166. (ZHU H X, HU L Z, DENG X L, et al. Molecular characterization and evolutionary analysis of DGAT1 gene family in three fabaceae plants[J]. Biotechnology Bulletin, 2011(10): 163-166.)[32]魏晨丹, 于继高, 滕佳, 等. 豆科全基因组DGAT基因家族的鉴定与进化分析[J]. 中国油料作物学报, 2020, 42(5): 807-817. (WEI C D, YU J G, TENG J, et al. Identification and evolutionary analysis of DGAT gene family in legumes[J]. Chinese Journal of Oil Crop Sciences, 2020, 42(5): 807-817.)[33]白玫, 吴鸿. 拟南芥TAG1基因对脂类合成调控作用的研究进展[J]. 植物学报, 2009, 44(6): 735-741. (BAI M, WU H. Recent progress in lipid biosynthesis regulated by TAG1 in Arabidopsis thaliana[J]. Bulletin of Botany, 2009, 44(6): 735-741.)[34]刘贵芹, 邵群, 黄荣峰, 等. 大豆DGAT基因家族的鉴定和表达分析[J]. 中国农学通报, 2013, 29(12): 55-61. (LIU G Q, SHAO Q, HUANG R F, et al. Characterization and expression analysis of DGAT gene family in soybean[J]. Chinese Agricultural Science Bulletin, 2013, 29(12): 55-61.)[35]张飞, 高慧玲, 刘宝玲, 等. 大豆GmDGAT3基因的鉴定及表达分析[J]. 山西农业科学, 2019, 47(4): 491-496. (ZHANG F, GAO H L, LIU B L, et al. Identification and expression analysis of soybean GmDGAT3 genes[J]. Journal of Shanxi Agricultural Sciences, 2019, 47(4): 491-496.)[36]晁毛妮, 胡喜贵, 张晋玉, 等. 大豆二酰甘油酰基转移酶基因GmDGAT1A启动子的克隆与功能分析[J]. 华北农学报, 2020, 35(4): 27-34. (CHAO M N, HU X G, ZHANG J Y, et al. Cloning and functional analysis of promoter of diacylglycerol acyltransferase gene GmDGAT1A in soybean[J]. Acta Agriculturae Boreali-Sinica, 2020, 35(4): 27-34.)[37]甘卓然, 石文茜, 黎永力, 等. 大豆生物钟基因GmLNK1/2、GmRVE4/8和GmTOC1 CRISPR/Cas9组织表达分析及敲除靶点的鉴定[J]. 作物学报, 2020, 46(8): 1291-1300. (GAN Z R, SHI W Q, LI Y L, et al. Identification of CRISPR/Cas9 knockout targets and tissue expression analysis of circadian clock genes GmLNK1/2, GmRVE4/8, and GmTOC1 in soybean[J]. Acta Agronomica Sinica, 2020, 46(8): 1291-1300.)[38]王计平, 张玲慧, 赵静, 等. 紫苏种子脂肪酸代谢及关键酶基因调控油脂合成规律的研究[J]. 中国粮油学报, 2016, 31(3): 5. (WANG J P, ZHANG L H, ZHAO J, et al. Regulation of controlling oil synthesis by fatty acid metabolism of Perilla seed and key enzyme gene[J]. Journal of the Chinese Cereals and Oils Association, 2016, 31(3): 5.)[39]CHENG Q, DONG L, GAO T, et al. The bHLH transcription factor GmPIB1 facilitates resistance to Phytophthora sojae in Glycine max[J]. Journal of Experimental Botany, 2018, 69(10): 2527-2541. [40]高宇, 陈莹, 孙岩, 等. 特色油料作物油莎豆CeDGAT1基因的鉴定及表达分析[J]. 山西农业科学, 2020, 48(6): 831-835, 841. (GAO Y, CHEN Y, SUN Y, et al. Identification and expression analysis of CeDGAT1 gene in Cyperus esculentus with a special oil crop[J]. Journal of Shanxi Agricultural Sciences, 2020, 48(6): 831-835, 841.)

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Last Update: 2022-08-10