通过CRISPR/Cas9技术突变BnMLO6基因提高甘蓝型油菜的抗病性
来源： 时间：2026-03-17 15:05
[1] 李利霞, 陈碧云, 闫贵欣, 高桂珍, 许鲲, 谢婷, 张付贵, 伍晓明. 中国油菜种质资源研究利用策略与进展. 植物遗传资源学报, 2020, 21:1-19. Li L X, Chen B Y, Yan G X, Gao G Z, Xu K, Xie T, Zhang F G, Wu X M. Proposed strategies and current progress of research and utilization of oilseed rape germplasm in China. J Plant Genet Resour, 2020, 21:1-19 (in Chinese with English abstract).[2] 刘成, 冯中朝, 肖唐华, 马晓敏, 周广生, 黄凤洪, 李加纳, 王汉中. 我国油菜产业发展现状、潜力及对策. 中国油料作物学报, 2019, 41:485-489. Liu C, Feng Z C, Xiao T H, Ma X M, Zhou G S, Huang F H, Li J N, Wang H Z. Development, potential and adaptation of Chinese rapeseed industry. Chin J Oil Crop Sci, 2019, 41:485-489 (in Chinese with English abstract).[3] 孙祥良, 王华弟, 曹奎荣, 朱金良. 油菜菌核病对油菜千粒重及产量的影响. 浙江农业科学, 2014, 11:1732-1733. Sun X L, Wang H D, Cao K R, Zhu J L. Effects of Sclerotinia sclerotinia on 1000-grain weight and yield of rapeseed. J Zhe jiang Agric Sci, 2014, 11:1732-1733 (in Chinese with English abstract).[4] 吴健, 周永明, 王幼平. 油菜与核盘菌互作分子机理研究进展. 中国油料作物学报, 2018, 40:721-729. Wu J, Zhou Y M, Wang Y P. Research progress on molecular mechanisms of Brassica napus-Sclerotinia sclerotiorum interaction. Chin J Oil Crop Sci, 2018, 40:721-729 (in Chinese with English abstract).[5] 杨清坡, 刘万才, 黄冲. 近10年油菜主要病虫害发生危害情况的统计和分析. 植物保护, 2018, 44(3):24-30. Yang Q P, Liu W C, Huang C. Statistics and analysis of oilseed rape losses caused by main diseases and insect pests in recent 10 years. Plant Prot, 2018, 44(3):24-30 (in Chinese with English abstract).[6] Gaetán S, Madia M. First report of canola powdery mildew caused by Erysiphe polygoni in Argentina. Plant Dis, 2004, 88:1163.
doi: 10.1094/PDIS.2004.88.10.1163Cpmid: 30795271[7] 邵登魁. 油菜抗白粉病鉴定及相关的生理生化特性研究. 甘肃农业大学硕士学位论文,甘肃兰州, 2006. Shao D K. Identification of Resistance to Erysiphe cruciferarum Junell and Study on Enzymes Associated with PM in Brassica Rape. MS Thesis of Gansu Agricultural University, Lanzhou, Gansu,China, 2006 (in Chinese with English abstract).[8] Tyagi S, Kumar R, Kumar V, Won S Y, Shukla P. Engineering disease resistant plants through CRISPR-Cas9 technology. GM Crop Food, 2021, 12:125-144.[9] 单奇伟, 高彩霞. 植物基因组编辑及衍生技术最新研究进展. 遗传, 2015, 37:953-973. Shan Q W, Gao C X. Research progress of genome editing and derivative technologies in plants. Hereditas, 2015, 37:953-973 (in Chinese with English abstract).[10] 刘耀光, 李构思, 张雅玲, 陈乐天. CRISPR/Cas植物基因组编辑技术研究进展. 华南农业大学学报, 2019, 40(5):38-49. Liu Y G, Li G S, Zhang Y L, Chen L T. Current advances on CRISPR/Cas genome editing technologies in plants. J South China Agric Univ, 2019, 40(5):38-49 (in Chinese with English abstract).[11] Cong L, Ran F A, Cox D, Lin S, Barretto R, Habib N, Hsu P D, Wu X, Jiang W, Marraffini L A, Zhang F. Multiplex genome engineering using CRISPR/Cas systems. Science, 2013, 339:819-823.
doi: 10.1126/science.1231143pmid: 23287718[12] Sorek R, Lawrence C M, Wiedenheft B. CRISPR-mediated adaptive immune systems in bacteria and archaea. Annu Rev Biochem, 2013, 82:237-266.
doi: 10.1146/biochem.2013.82.issue-1[13] Mao Y, Zhang H, Xu N, Zhang B, Gou F, Zhu J K. Application of the CRISPR-Cas system for efficient genome engineering in plants. Mol Plant, 2013, 6:2008-2011.
doi: 10.1093/mp/sst121[14] Xie K, Yang Y. RNA-guided genome editing in plants using a CRISPR-Cas system. Mol Plant, 2013, 6:1975-1983.
doi: 10.1093/mp/sst119[15] Upadhyay S K, Kumar J, Alok A, Tuli R. RNA-guided genome editing for target gene mutations in wheat. G3: Gen Genom Genet, 2013, 3:2233-2238.[16] Liang Z, Zhang K, Chen K, Gao C. Targeted mutagenesis in Zea mays using TALENs and the CRISPR/Cas system. J Genet Genomics, 2014, 41:63-68.
doi: 10.1016/j.jgg.2013.12.001pmid: 24576457[17] Gao J, Wang G, Ma S, Xie X, Wu X, Zhang X, Wu Y, Zhao P, Xia Q. CRISPR/Cas9-mediated targeted mutagenesis in Nicotiana tabacum. Plant Mol Biol, 2015, 87:99-110.
doi: 10.1007/s11103-014-0263-0[18] Soyk S, Müller N A, Park S J, Schmalenbach I, Jiang K, Hayama R, Zhang L, Van Eck J, Jiménez-Gómez J M, Lippman Z B. Variation in the flowering gene SELF PRUNING 5G promotes day-neutrality and early yield in tomato. Nat Genet, 2017, 49:162-168.
doi: 10.1038/ng.3733[19] Braatz J, Harloff H J, Mascher M, Stein N, Himmelbach A, Jung C. CRISPR-Cas9 targeted mutagenesis leads to simultaneous modification of different homoeologous gene copies in polyploid oilseed rape (Brassica napus). Plant Physiol, 2017, 174:935-942.
doi: 10.1104/pp.17.00426[20] Jørgensen I H. Discovery, characterization and exploitation of MLO powdery mildew resistance in barley. Euphytica, 1992, 63:141-152.
doi: 10.1007/BF00023919[21] Devoto A, Hartmann H A, Piffanelli P, Elliott C, Simmons C, Taramino G, Goh C S, Cohen F E, Emerson B C, Schulze-Lefert P, Panstruga R. Molecular phylogeny and evolution of the plant- specific seven-transmembrane MLO family. J Mol Evol, 2003, 56:77-88.
doi: 10.1007/s00239-002-2382-5[22] Liu Q, Zhu H. Molecular evolution of the MLO gene family in Oryza sativa and their functional divergence. Gene, 2008, 409:1-10.
doi: 10.1016/j.gene.2007.10.031[23] Konishi S, Sasakuma T, Sasanuma T. Identification of novel MLO family members in wheat and their genetic characterization. Genes Genet Syst, 2010, 85:167-175.
pmid: 21041976[24] Zhou S J, Jing Z, Shi J L. Genome-wide identification, characterization, and expression analysis of the MLO gene family in Cucumis sativus. Genet Mol Res, 2013, 12:6565-6578.
doi: 10.4238/2013.December.11.8pmid: 24391003[25] Wolter M, Hollricher K, Salamini F, Schulze-Lefert P. The mlo resistance alleles to powdery mildew infection in barley trigger a developmentally controlled defence mimic phenotype. Mol Gen Genet, 1993, 239:122-128.
doi: 10.1007/BF00281610[26] Ropenack E V, Parr A, Schulze-Lefert P. Structural analyses and dynamics of soluble and cell wall-bound phenolics in a broad spectrum resistance to the powdery mildew fungus in barley. J Biol Chem, 1998, 273:9013-9022.
doi: 10.1074/jbc.273.15.9013[27] Piffanelli P, Zhou F, Casais C, Orme J, Jarosch B, Schaffrath U, Collins N C, Panstruga R, Schulze-Lefert P. The barley MLO modulator of defense and cell death is responsive to biotic and abiotic stress stimuli. Plant Physiol, 2002, 129:1076-1085.
pmid: 12114562[28] Kuhn H, Lorek J, Kwaaitaal M, Becker K, Micali C, Ver Loren van Themaat E, Bednarek P, Raaymakers T M, Appiano M, Bai Y, Meldau D, Baum S, Conrath U, Feussner I, Panstruga R. Key components of different plant defense pathways are dispensable for powdery mildew resistance of the Arabidopsis mlo2 mlo6 mlo12 triple mutant. Front Plant Sci, 2017, 8:1006.
doi: 10.3389/fpls.2017.01006[29] 贾云飞, 张国海, 刘崇怀, 樊秀彩, 姜建福, 孙海生, 张颖. Mlo基因在葡萄抗白腐病中作用的研究. 植物生理学报, 2017, 53:1649-1658. Jia Y F, Zhang G H, Liu C H, Fan X C, Jiang J F, Sun H S, Zhang Y. Study on the function of resistance to white rot of Mlo genes in grapevine. Plant Physiol J, 2017, 53:1649-1658 (in Chinese with English abstract).[30] Acevedo-Garcia J, Gruner K, Reinstädler A, Kemen A, Kemen E, Cao L, Takken F L W, Reitz M U, Schäfer P, O’Connell R J, Kusch S, Kuhn H, Panstruga R. The powdery mildew-resistant Arabidopsis mlo2 mlo6 mlo12 triple mutant displays altered infection phenotypes with diverse types of phytopathogens. Sci Rep, 2017, 7:9319.
doi: 10.1038/s41598-017-07188-7pmid: 28839137[31] McGrann G R, Stavrinides A, Russell J, Corbitt M M, Booth A, Chartrain L, Thomas W T, Brown J K. A trade off between mlo resistance to powdery mildew and increased susceptibility of barley to a newly important disease, Ramularia leaf spot. J Exp Bot, 2014, 65:1025-1037.
doi: 10.1093/jxb/ert452pmid: 24399175[32] Shi J, Wan H, Zai W, Xiong Z, Wu W. Phylogenetic relationship of plant MLO genes and transcriptional response of MLO genes to Ralstonia solanacearum in tomato. Genes, 2020, 11:487.
doi: 10.3390/genes11050487[33] Ma X, Zhang Q, Zhu Q, Liu W, Chen Y, Qiu R, Wang B, Yang Z, Li H, Lin Y, Xie Y, Shen R, Chen S, Wang Z, Chen Y, Guo J, Chen L, Zhao X, Dong Z, Liu Y G. A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants. Mol Plant, 2015, 8:1274-1284.
doi: 10.1016/j.molp.2015.04.007[34] Li C, Hao M, Wang W, Wang H, Chen F, Chu W, Zhang B, Mei D, Cheng H, Hu Q. An efficient CRISPR/Cas9 platform for rapidly generating simultaneous mutagenesis of multiple gene homeologs in allotetraploid oilseed rape. Front Plant Sci, 2018, 9:442.
doi: 10.3389/fpls.2018.00442[35] Luna E, Pastor V, Robert J, Flors V, Mauch-Mani B, Ton J. Callose deposition: a multifaceted plant defense response. Mol Plant Microbe Interact, 2011, 24:183-193.
doi: 10.1094/MPMI-07-10-0149[36] 何烈干, 宋来强, 汤洁, 周银生, 马辉刚. 油菜菌核病抗性鉴定方法比较及抗病种质资源的筛选. 江苏农业科学, 2018, 46(18):90-93. He L G, Song L Q, Tang J, Zhou Y S, Ma H G. Comparison of identification methods for resistance to Sclerotinia sclerotiorum and screening of resistant materials of rapeseed. Jiangsu Agric Sci, 2018, 46(18):90-93 (in Chinese).[37] Zaidi S S, Mukhtar M S, Mansoor S. Genome editing: targeting susceptibility genes for plant disease resistance. Trends Biotechnol, 2018, 36:898-906.
doi: 10.1016/j.tibtech.2018.04.005[38] Sun Q, Lin L, Liu D, Wu D, Fang Y, Wu J, Wang Y. CRISPR/Cas9-mediated multiplex genome editing of the BnWRKY11 and BnWRKY70 genes in Brassica napus L. Int J Mol Sci, 2018, 19:2716.
doi: 10.3390/ijms19092716[39] Naumann M, Somerville S, Voigt C. Differences in early callose deposition during adapted and non-adapted powdery mildew infection of resistant Arabidopsis lines. Plant Signal Behav, 2013, 8:e24408.[40] Bari R, Jones J D G. Role of plant hormones in plant defence responses. Plant Mol Biol, 2009, 69:473-488.
doi: 10.1007/s11103-008-9435-0
doi: 10.1094/PDIS.2004.88.10.1163Cpmid: 30795271[7] 邵登魁. 油菜抗白粉病鉴定及相关的生理生化特性研究. 甘肃农业大学硕士学位论文,甘肃兰州, 2006. Shao D K. Identification of Resistance to Erysiphe cruciferarum Junell and Study on Enzymes Associated with PM in Brassica Rape. MS Thesis of Gansu Agricultural University, Lanzhou, Gansu,China, 2006 (in Chinese with English abstract).[8] Tyagi S, Kumar R, Kumar V, Won S Y, Shukla P. Engineering disease resistant plants through CRISPR-Cas9 technology. GM Crop Food, 2021, 12:125-144.[9] 单奇伟, 高彩霞. 植物基因组编辑及衍生技术最新研究进展. 遗传, 2015, 37:953-973. Shan Q W, Gao C X. Research progress of genome editing and derivative technologies in plants. Hereditas, 2015, 37:953-973 (in Chinese with English abstract).[10] 刘耀光, 李构思, 张雅玲, 陈乐天. CRISPR/Cas植物基因组编辑技术研究进展. 华南农业大学学报, 2019, 40(5):38-49. Liu Y G, Li G S, Zhang Y L, Chen L T. Current advances on CRISPR/Cas genome editing technologies in plants. J South China Agric Univ, 2019, 40(5):38-49 (in Chinese with English abstract).[11] Cong L, Ran F A, Cox D, Lin S, Barretto R, Habib N, Hsu P D, Wu X, Jiang W, Marraffini L A, Zhang F. Multiplex genome engineering using CRISPR/Cas systems. Science, 2013, 339:819-823.
doi: 10.1126/science.1231143pmid: 23287718[12] Sorek R, Lawrence C M, Wiedenheft B. CRISPR-mediated adaptive immune systems in bacteria and archaea. Annu Rev Biochem, 2013, 82:237-266.
doi: 10.1146/biochem.2013.82.issue-1[13] Mao Y, Zhang H, Xu N, Zhang B, Gou F, Zhu J K. Application of the CRISPR-Cas system for efficient genome engineering in plants. Mol Plant, 2013, 6:2008-2011.
doi: 10.1093/mp/sst121[14] Xie K, Yang Y. RNA-guided genome editing in plants using a CRISPR-Cas system. Mol Plant, 2013, 6:1975-1983.
doi: 10.1093/mp/sst119[15] Upadhyay S K, Kumar J, Alok A, Tuli R. RNA-guided genome editing for target gene mutations in wheat. G3: Gen Genom Genet, 2013, 3:2233-2238.[16] Liang Z, Zhang K, Chen K, Gao C. Targeted mutagenesis in Zea mays using TALENs and the CRISPR/Cas system. J Genet Genomics, 2014, 41:63-68.
doi: 10.1016/j.jgg.2013.12.001pmid: 24576457[17] Gao J, Wang G, Ma S, Xie X, Wu X, Zhang X, Wu Y, Zhao P, Xia Q. CRISPR/Cas9-mediated targeted mutagenesis in Nicotiana tabacum. Plant Mol Biol, 2015, 87:99-110.
doi: 10.1007/s11103-014-0263-0[18] Soyk S, Müller N A, Park S J, Schmalenbach I, Jiang K, Hayama R, Zhang L, Van Eck J, Jiménez-Gómez J M, Lippman Z B. Variation in the flowering gene SELF PRUNING 5G promotes day-neutrality and early yield in tomato. Nat Genet, 2017, 49:162-168.
doi: 10.1038/ng.3733[19] Braatz J, Harloff H J, Mascher M, Stein N, Himmelbach A, Jung C. CRISPR-Cas9 targeted mutagenesis leads to simultaneous modification of different homoeologous gene copies in polyploid oilseed rape (Brassica napus). Plant Physiol, 2017, 174:935-942.
doi: 10.1104/pp.17.00426[20] Jørgensen I H. Discovery, characterization and exploitation of MLO powdery mildew resistance in barley. Euphytica, 1992, 63:141-152.
doi: 10.1007/BF00023919[21] Devoto A, Hartmann H A, Piffanelli P, Elliott C, Simmons C, Taramino G, Goh C S, Cohen F E, Emerson B C, Schulze-Lefert P, Panstruga R. Molecular phylogeny and evolution of the plant- specific seven-transmembrane MLO family. J Mol Evol, 2003, 56:77-88.
doi: 10.1007/s00239-002-2382-5[22] Liu Q, Zhu H. Molecular evolution of the MLO gene family in Oryza sativa and their functional divergence. Gene, 2008, 409:1-10.
doi: 10.1016/j.gene.2007.10.031[23] Konishi S, Sasakuma T, Sasanuma T. Identification of novel MLO family members in wheat and their genetic characterization. Genes Genet Syst, 2010, 85:167-175.
pmid: 21041976[24] Zhou S J, Jing Z, Shi J L. Genome-wide identification, characterization, and expression analysis of the MLO gene family in Cucumis sativus. Genet Mol Res, 2013, 12:6565-6578.
doi: 10.4238/2013.December.11.8pmid: 24391003[25] Wolter M, Hollricher K, Salamini F, Schulze-Lefert P. The mlo resistance alleles to powdery mildew infection in barley trigger a developmentally controlled defence mimic phenotype. Mol Gen Genet, 1993, 239:122-128.
doi: 10.1007/BF00281610[26] Ropenack E V, Parr A, Schulze-Lefert P. Structural analyses and dynamics of soluble and cell wall-bound phenolics in a broad spectrum resistance to the powdery mildew fungus in barley. J Biol Chem, 1998, 273:9013-9022.
doi: 10.1074/jbc.273.15.9013[27] Piffanelli P, Zhou F, Casais C, Orme J, Jarosch B, Schaffrath U, Collins N C, Panstruga R, Schulze-Lefert P. The barley MLO modulator of defense and cell death is responsive to biotic and abiotic stress stimuli. Plant Physiol, 2002, 129:1076-1085.
pmid: 12114562[28] Kuhn H, Lorek J, Kwaaitaal M, Becker K, Micali C, Ver Loren van Themaat E, Bednarek P, Raaymakers T M, Appiano M, Bai Y, Meldau D, Baum S, Conrath U, Feussner I, Panstruga R. Key components of different plant defense pathways are dispensable for powdery mildew resistance of the Arabidopsis mlo2 mlo6 mlo12 triple mutant. Front Plant Sci, 2017, 8:1006.
doi: 10.3389/fpls.2017.01006[29] 贾云飞, 张国海, 刘崇怀, 樊秀彩, 姜建福, 孙海生, 张颖. Mlo基因在葡萄抗白腐病中作用的研究. 植物生理学报, 2017, 53:1649-1658. Jia Y F, Zhang G H, Liu C H, Fan X C, Jiang J F, Sun H S, Zhang Y. Study on the function of resistance to white rot of Mlo genes in grapevine. Plant Physiol J, 2017, 53:1649-1658 (in Chinese with English abstract).[30] Acevedo-Garcia J, Gruner K, Reinstädler A, Kemen A, Kemen E, Cao L, Takken F L W, Reitz M U, Schäfer P, O’Connell R J, Kusch S, Kuhn H, Panstruga R. The powdery mildew-resistant Arabidopsis mlo2 mlo6 mlo12 triple mutant displays altered infection phenotypes with diverse types of phytopathogens. Sci Rep, 2017, 7:9319.
doi: 10.1038/s41598-017-07188-7pmid: 28839137[31] McGrann G R, Stavrinides A, Russell J, Corbitt M M, Booth A, Chartrain L, Thomas W T, Brown J K. A trade off between mlo resistance to powdery mildew and increased susceptibility of barley to a newly important disease, Ramularia leaf spot. J Exp Bot, 2014, 65:1025-1037.
doi: 10.1093/jxb/ert452pmid: 24399175[32] Shi J, Wan H, Zai W, Xiong Z, Wu W. Phylogenetic relationship of plant MLO genes and transcriptional response of MLO genes to Ralstonia solanacearum in tomato. Genes, 2020, 11:487.
doi: 10.3390/genes11050487[33] Ma X, Zhang Q, Zhu Q, Liu W, Chen Y, Qiu R, Wang B, Yang Z, Li H, Lin Y, Xie Y, Shen R, Chen S, Wang Z, Chen Y, Guo J, Chen L, Zhao X, Dong Z, Liu Y G. A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in monocot and dicot plants. Mol Plant, 2015, 8:1274-1284.
doi: 10.1016/j.molp.2015.04.007[34] Li C, Hao M, Wang W, Wang H, Chen F, Chu W, Zhang B, Mei D, Cheng H, Hu Q. An efficient CRISPR/Cas9 platform for rapidly generating simultaneous mutagenesis of multiple gene homeologs in allotetraploid oilseed rape. Front Plant Sci, 2018, 9:442.
doi: 10.3389/fpls.2018.00442[35] Luna E, Pastor V, Robert J, Flors V, Mauch-Mani B, Ton J. Callose deposition: a multifaceted plant defense response. Mol Plant Microbe Interact, 2011, 24:183-193.
doi: 10.1094/MPMI-07-10-0149[36] 何烈干, 宋来强, 汤洁, 周银生, 马辉刚. 油菜菌核病抗性鉴定方法比较及抗病种质资源的筛选. 江苏农业科学, 2018, 46(18):90-93. He L G, Song L Q, Tang J, Zhou Y S, Ma H G. Comparison of identification methods for resistance to Sclerotinia sclerotiorum and screening of resistant materials of rapeseed. Jiangsu Agric Sci, 2018, 46(18):90-93 (in Chinese).[37] Zaidi S S, Mukhtar M S, Mansoor S. Genome editing: targeting susceptibility genes for plant disease resistance. Trends Biotechnol, 2018, 36:898-906.
doi: 10.1016/j.tibtech.2018.04.005[38] Sun Q, Lin L, Liu D, Wu D, Fang Y, Wu J, Wang Y. CRISPR/Cas9-mediated multiplex genome editing of the BnWRKY11 and BnWRKY70 genes in Brassica napus L. Int J Mol Sci, 2018, 19:2716.
doi: 10.3390/ijms19092716[39] Naumann M, Somerville S, Voigt C. Differences in early callose deposition during adapted and non-adapted powdery mildew infection of resistant Arabidopsis lines. Plant Signal Behav, 2013, 8:e24408.[40] Bari R, Jones J D G. Role of plant hormones in plant defence responses. Plant Mol Biol, 2009, 69:473-488.
doi: 10.1007/s11103-008-9435-0
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