外源注射褪黑素对小麦抵抗叶锈菌侵染的影响

doi:10.7668/hbnxb.20192998

摘要/Abstract

摘要：

为了探究褪黑素在小麦与叶锈菌互作中的作用,以小麦品种洛夫林10(简称L10)与叶锈菌生理小种260组成不亲和组合为研究对象,通过外源注射甲基紫精(甲基紫精作为一种氧化剂诱发产生超氧阴离子,可以有效增加活性氧的含量),诱导产生活性氧,利用褪黑素清除活性氧的能力确定褪黑素最佳作用浓度;然后在小麦幼苗叶片中注射褪黑素并接种叶锈菌生理小种260,通过DAB染色观察H₂O₂含量变化;通过Rohringer染色检测HR面积;通过测定小麦过氧化物酶和过氧化氢酶活性,探究外源注射褪黑素对小麦抗氧化能力的影响;通过以上研究以明确褪黑素在小麦与叶锈菌互作中的作用。结果表明:外源注射甲基紫精引起的活性氧增加,其清除最适的褪黑素注射浓度为10 μmol/L。对不亲和组合的DAB染色结果显示,注射10 μmol/L褪黑素后,叶锈菌侵染诱发的小麦叶片H₂O₂积累量少于对照组,表明褪黑素参与了H₂O₂清除作用; Rohringer染色结果表明,经过外源性褪黑素处理后小麦HR细胞面积减小,有效增强了对叶锈菌的抗性。此外,外源注射褪黑素提高了抗氧化酶POD和CAT的活性,表明小麦的抗氧化能力获得显著提升。试验结果表明,外源注射褪黑素参与了小麦抗叶锈菌过程中活性氧的清除,提高了小麦的抗病能力。

关键词: 小麦, 褪黑素, 活性氧, DAB, HR, 抗氧化酶

Abstract:

In order to explore the role of melatonin in the interaction between wheat and leaf rust,we studied the incompatible combination between wheat variety Lovrin 10(hereinafter referred to as L10)and physiological race 260 of leaf rust. In this experiment,reactive oxygen species were induced by exogenous injection of Methyl viologen (Methy lviologen(MV)as an oxidant induces the production of superoxide anion,which can effectively increase the content of reactive oxygen species),and the best concentration of melatonin was determined by using the ability of melatonin to scavenge reactive oxygen species;then,melatonin was injected into the leaves of wheat seedlings and inoculated with leaf rust race 260,and the changes of H₂O₂ content were observed by DAB staining. The HR area was detected by Rohringer staining. The effects of exogenous melatonin injection on the antioxidant capacity of wheat were investigated by measuring the peroxidase(POD)and catalase(CAT)activities of wheat;through the above research,the role of melatonin in the interaction between wheat and leaf rust was clarified. The results showed that the reactive oxygen species was increased by exogenous injection of methyl viologen,and the optimum injection concentration of melatonin was 10 μmol/L. The results of DAB staining of incompatible combinations showed that after injection of 10 μmol/L melatonin,the accumulation of H₂O₂ induced by leaf rust infection in wheat leaves was less than that in control group,which indicated that melatonin participated in H₂O₂ scavenging. Rohringer staining showed that the area of wheat HR cells decreased after exogenous melatonin treatment,which effectively enhanced the resistance to leaf rust. In addition,exogenous injection of melatonin increased the activities of antioxidant enzymes POD and CAT,which indicated that the antioxidant capacity of wheat was significantly improved. The results showed that exogenous injection of melatonin was involved in the removal of reactive oxygen species in the process of wheat resistance to leaf rust,and improved the disease resistance of wheat.

Key words: Wheat, Melatonin, Reactive oxygen species, DAB, HR, Antioxidant enzyme

王硕, 赵港伊, 史天乐, 吴思凡, 闫倩颖, 韩胜芳, 王冬梅. 外源注射褪黑素对小麦抵抗叶锈菌侵染的影响[J]. 华北农学报, 2022, 37(5): 181-186. doi: 10.7668/hbnxb.20192998.

WANG Shuo, ZHAO Gangyi, SHI Tianle, WU Sifan, YAN Qianying, HAN Shengfang, WANG Dongmei. Effect of Exogenous Melatonin Injection on Resistance of Wheat to Leaf Rust[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(5): 181-186. doi: 10.7668/hbnxb.20192998.

http://www.hbnxb.net/CN/Y2022/V37/I5/181

图/表 5

图1 不同浓度褪黑素对甲基紫精诱发产生H₂O₂的清除作用

Fig.1 Scavenging effect of melatonin with different concentrations on H₂O₂ induced by methyl viologen

图2 外源注射褪黑素小麦叶片的DAB染色观察

Fig.2 DAB staining observation of wheat leaves injected with melatonin

图3 外源注射褪黑素的小麦HR细胞染色观察

Fig.3 Observation on HR cell staining of wheat injected with melatonin

图4 外源注射褪黑素小麦HR细胞面积不同小写字母表示差异显著(P<0.05)。图5同。

Fig.4 Statistical results of HR cell area of wheat injected with melatonin Different lowercase letters indicate significant differences (P<0.05).The same as Fig.5.

图5 注射褪黑素对小麦叶片POD(A)、CAT(B)活性的影响

Fig.5 Effects of melatonin injection on POD(A)and CAT(B)activities in wheat leaves

参考文献 23

[1]	夏小双. 中国小麦叶锈菌越夏区划及群体遗传结构分析[D]. 北京: 中国农业科学院, 2021.
	Xia X S. Over-summering regionalization and population genetic structure of Puccinia triticina in China[D]. Beijing: Chinese Academy of Agricultural Sciences, 2021.
[2]	齐悦, 吕峻元, 张悦, 韦杰, 张娜, 杨文香, 刘大群. 小麦叶锈菌效应蛋白Pt18906激发TcLr27+31的双层防御反应[J]. 中国农业科学, 2020, 53(12):2371-2384. doi: 10.3864/j.issn.0578-1752.2020.12.006. doi: 10.3864/j.issn.0578-1752.2020.12.006
	Qi Y, Lü J Y, Zhang Y, Wei J, Zhang N, Yang W X, Liu D Q. Puccinia triticina effector protein Pt18906 triggered two-layer defense reaction in TcLr27+31[J]. Scientia Agricultura Sinica, 2020, 53(12):2371-2384.
[3]	李彤彤, 宋姗姗, 王琦, 侯春燕, 王冬梅. 小麦与叶锈菌互作过程中精氨酸脱羧酶基因TaADC的功能研究[J]. 农业生物技术学报, 2020, 28(8):1343-1351. doi: 10.3969/j.issn.1674-7968.2020.08.002. doi: 10.3969/j.issn.1674-7968.2020.08.002
	Li T T, Song S S, Wang Q, Hou C Y, Wang D M. Study on the function of arginine decarboxylase gene TaADC in the interaction between wheat(Triticum aestivum)and Puccinia triticina[J]. Journal of Agricultural Biotechnology, 2020, 28(8):1343-1351.
[4]	张融雪, 孙玥, 苏京平, 王胜军, 佟卉, 刘燕清, 孙林静. 植物褪黑素研究进展[J]. 生物技术进展, 2021, 11(3):297-303. doi: 10.19586/j.2095-2341.2020.0148. doi: 10.19586/j.2095-2341.2020.0148
	Zhang R X, Sun Y, Su J P, Wang S J, Tong H, Liu Y Q, Sun L J. Research progress on plant melatonin[J]. Current Biotechnology, 2021, 11(3):297-303.
[5]	Aghdam M S, Fard J R. Melatonin treatment attenuates postharvest decay and maintains nutritional quality of strawberry fruits(Fragaria anannasa cv. Selva)by enhancing GABA shunt activity[J]. Food Chemistry, 2017, 221:1650-1657. doi: 10.1016/j.foodchem.2016.10.123. doi: 10.1016/j.foodchem.2016.10.123 URL
[6]	Sharif R, Xie C, Zhang H Q, Arnao M B, Ali M, Ali Q, Muhammad I, Shalmani A, Nawaz M A, Chen P, Li Y H. Melatonin and its effects on plant systems[J]. Molecules, 2018, 23(9):2352. doi: 10.3390/molecules23092352. doi: 10.3390/molecules23092352 URL
[7]	Moustafa-Farag M, Almoneafy A, Mahmoud A, Elkelish A, Arnao M B, Li L F, Ai S Y. Melatonin and its protective role against biotic stress impacts on plants[J]. Biomolecules, 2019, 10(1):54. doi: 10.3390/biom10010054. doi: 10.3390/biom10010054 URL
[8]	王光涛, 冯素伟, 丁位华, 胡喜贵, 文昭普, 茹振钢. 根际pH值对冬小麦叶片抗氧化酶活性和内源激素含量的影响[J]. 江苏农业科学, 2021, 49(10):71-75. doi: 10.15889/j.issn.1002-1302.2021.10.013. doi: 10.15889/j.issn.1002-1302.2021.10.013
	Wang G T, Feng S W, Ding W H, Hu X G, Wen Z P, Ru Z G. Effects of rhizosphere pH value on antioxidant enzyme activity and endogenous hormone content in winter wheat leaves[J]. Jiangsu Agricultural Sciences, 2021, 49(10):71-75.
[9]	Back K. Melatonin metabolism,signaling and possible roles in plants[J]. The Plant Journal, 2021, 105(2):376-391. doi: 10.1111/tpj.14915. doi: 10.1111/tpj.14915 URL
[10]	Hans T C, Zhang Z G, Wei Y D, Collinge D B. Subcellular localization of H₂O₂ in plants. H₂O₂ accumulation in papillae and hypersensitive response during the barley-powdery mildew interaction[J]. The Plant Journal, 1997, 11(6):1187-1194.doi: 10.1046/j.1365-313x.1997.11061187.x. doi: 10.1046/j.1365-313x.1997.11061187.x URL
[11]	Rohringer R. Calcofluor:an optical brightener for fluorescence microscopy of fungal plant parasites in leaves[J]. Phytopathology, 1977, 77(6):808. doi: 10.1094/phyto-67-808. doi: 10.1094/phyto-67-808
[12]	邹琦. 植物生理学实验指导[M]. 北京: 中国农业出版社, 2000.
	Zou Q. Experimental guidance of plant physiology[M]. Beijing: Higher Education Press, 2000.
[13]	Zang M H, Zhao Y C, Gao L C, Zhong F Y, Qin Z H, Tong R Y, Ai L L, Petersen L, Yan Y, Gao Y, Zhu C S, Pu J. The circadian nuclear receptor ROR α negatively regulates cerebral ischemia-reperfusion injury and mediates the neuroprotective effects of melatonin[J]. Biochim Biophys Acta Mol Basis Dis, 2020, 1866(11): 165890. doi: 10.1016/j.bbadis.2020.165890. doi: 10.1016/j.bbadis.2020.165890 URL
[14]	Reiter R J. Pineal melatonin:Cell biology of its synthesis and of its physiological interactions[J]. Endocrine Reviews, 1991, 12(2):151-180. doi: 10.1210/edrv-12-2-151. doi: 10.1210/edrv-12-2-151 pmid: 1649044
[15]	陈贤, 杨勇, 刘凤权. 植物褪黑素的研究进展[J]. 江苏农业科学, 2020, 48(24):17-24. doi: 10.15889/j.issn.1002-1302.2020.24.003. doi: 10.15889/j.issn.1002-1302.2020.24.003
	Chen X, Yang Y, Liu F Q. Research progress of plant melatonin[J]. Jiangsu Agricultural Sciences, 2020, 48(24):17-24.
[16]	Yin L H, Wang P, Li M J, Ke X W, Li C Y, Liang D, Wu S, Ma X L, Li C, Zou Y J, Ma F W. Exogenous melatonin improves Malus resistance to Marssonina apple blotch[J]. Journal of Pineal Research, 2013, 54(4):426-434. doi: 10.1111/jpi.12038. doi: 10.1111/jpi.12038 URL
[17]	辛丹丹, 司金金, 寇莉萍. 黄瓜采后外源褪黑素处理提高品质和延缓衰老的研究[J]. 园艺学报, 2017, 44(5):891-901. doi: 10.16420/j.issn.0513-353x.2016-0888. doi: 10.16420/j.issn.0513-353x.2016-0888
	Xin D D, Si J J, Kou L P. Postharvest exogenous melatonin enhances quality and delays the senescence of cucumber[J]. Acta Horticulturae Sinica, 2017, 44(5):891-901.
[18]	Liu C X, Chen L L, Zhao R R, Li R, Zhang S J, Yu W Q, Sheng J P, Shen L. Melatonin induces disease resistance to Botrytis cinerea in tomato fruit by activating jasmonic acid signaling pathway[J]. Journal of Agricultural and Food Chemistry, 2019, 67(22):6116-6124. doi: 10.1021/acs.jafc.9b00058. doi: 10.1021/acs.jafc.9b00058 URL
[19]	Mandal M K, Suren H, Ward B, Boroujerdi A, Kousik C. Differential roles of melatonin in plant-host resistance and pathogen suppression in cucurbits[J]. Journal of Pineal Research, 2018, 65(3):e12505. doi: 10.1111/jpi.12505. doi: 10.1111/jpi.12505
[20]	Lin Y L, Fan L Q, Xia X H, Wang Z K, Yin Y P, Cheng Y L, Li Z G. Melatonin decreases resistance to postharvest green mold on Citrus fruit by scavenging defense-related reactive oxygen species[J]. Postharvest Biology and Technology, 2019, 153:21-30. doi: 10.1016/j.postharvbio.2019.03.016. doi: 10.1016/j.postharvbio.2019.03.016 URL
[21]	臧华伟. 外源褪黑素和硒对番茄果实采后灰霉病的控制作用及机理[D]. 合肥: 安徽农业大学, 2020.
	Zang H W. Exogenous melatonin and selenium induce disease resistance to postharvest gray mold in tomato fruit[D]. Hefei: Anhui Agricultural University, 2020.
[22]	郭博铖, 柯希望, 高尚雨, 于昕卉, 孙启明, 左豫虎. 褪黑素诱导小豆抗锈病机理的初步研究[J]. 植物保护, 2020, 46(1):145-150,156. doi: 10.16688/j.zwbh.2018521. doi: 10.16688/j.zwbh.2018521
	Guo B C, Ke X W, Gao S Y, Yu X H, Sun Q M, Zuo Y H. A preliminary study on the mechanisms of melatonin-induced rust resistance of adzuki bean[J]. Plant Protection, 2020, 46(1):145-150,156.
[23]	Wei J, Li D X, Zhang J R, Shan C, Rengel Z, Song Z B, Chen Q. Phytomelatonin receptor PMTR1-mediated signaling regulates stomatal closure in Arabidopsis thaliana[J]. Journal of Pineal Research, 2018, 65(2):e12500. doi: 10.1111/jpi.12500. doi: 10.1111/jpi.12500 URL

选择文件类型/文献管理软件名称

选择包含的内容