外源褪黑素调控棉花枯萎病抗性研究

doi:10.13560/j.cnki.biotech.bull.1985.2022-0419

摘要/Abstract

摘要：

为探究外源褪黑素对棉花抗枯萎病的影响及作用机理，以海岛棉新海14号为材料，于叶面喷施不同浓度褪黑素（0、10、25、50、75、100 μmol/L）后接种棉花枯萎病菌，对枯萎病菌侵染后棉花幼苗进行抗病性鉴定、抗病相关酶活性及基因表达分析。结果表明，外源褪黑素对棉花枯萎病菌的生长无抑制作用，10-100 μmol/L褪黑素均能在一定程度上提高棉花对枯萎病的抗性，其中50 μmol/L褪黑素处理效果最好。与对照相比，50 μmol/L褪黑素预处理能有效减少接菌后棉花叶片中的过氧化氢（H₂O₂）含量和超氧阴离子（O₂^·-）的产生速率，增强过氧化物酶（POD）、过氧化氢酶（CAT）、超氧化物歧化酶（SOD）、抗坏血酸过氧化物酶（APX）、几丁质酶（CHT）、β-1,3葡聚糖酶（GLU）、多酚氧化酶（PPO）、苯丙氨酸解氨酶（PAL）活性，并显著提高木质素代谢途径相关基因（GbPAL、Gb4CL和GbCAD）和类黄酮代谢途径关键基因（GbCHI、GbDFR和GbTT7）的表达量。表明50 μmol/L 的褪黑素处理能提高接菌后棉花抗氧化能力，增强防御酶活性，调控木质素、类黄酮代谢途径相关基因的表达，从而提高棉花对枯萎病的抗性。

关键词: 褪黑素, 枯萎病, 棉花, 抗氧化酶, 抗病性

Abstract:

To investigate the effect and mechanism of exogenous melatonin on cotton resistance against Fusarium wilt, Gossypium barbadense Xinhai 14 treated with foliar application of various concentrations of melatonin（0, 10, 25, 50, 75 and 100 μmol/L）was used to identify disease resistance, to identify the activity of disease resistance related enzymes, and to analyze gene expression in cotton seedlings after inoculation with Fusarium oxysporum f.sp. vasinfectum（Fov）. The results showed that exogenous melatonin did not inhibit the growth of cotton blight, and they 10-100 μmol/L all increased the cotton resistance to Fov to some extent, the best with 50 μmol/L melatonin. Compared with control, 50 μmol/L melatonin pretreatment reduced the hydrogen peroxide（H₂O₂）content and superoxide anion（O₂^·-）production rate in the cotton leaves after inoculation, and enhanced the expressions of peroxidase（POD）, catalase（CAT）, superoxide dismutase（SOD）, ascorbate peroxidase（APX）, chitinase（CHT）, β-1,3 glucanase（GLU）, polyphenol oxidase（PPO）, phenylalanine ammonia lyase（PAL）activities, and significantly increased the expressions of key genes of the lignin synthesis pathway（GbPAL, Gb4CL and GbCAD）and key genes of the flavonoid synthesis pathway（GbCHI, GbDFR and GbTT7）. This showed that 50 μmol/L melatonin treatment may improve the antioxidant capacity of cotton after inoculation, enhance the activities of defense enzymes, regulate the expressions of genes related to the lignin and flavonoid synthesis pathways, and thus improve the resistance of cotton to Fusarium wilt.

Key words: melatonin, Fusarium wilt, cotton, antioxidant enzymes, disease resistance

朱金成, 杨洋, 娄慧, 张薇. 外源褪黑素调控棉花枯萎病抗性研究[J]. 生物技术通报, 2023, 39(1): 243-252.

ZHU Jin-cheng, YANG Yang, LOU Hui, ZHANG Wei. Regulation of Fusarium wilt Resistance in Cotton by Exogenous Melatonin[J]. Biotechnology Bulletin, 2023, 39(1): 243-252.

图/表 6

表1 荧光定量PCR引物序列

Table 1 Primer sequences for quantitative PCR

引物名称Primer name	引物序列Primer sequence（5'-3'）
引物名称Primer name	正向Forward（F）	反向Reverse（R）
GbPAL-F/R	CCAATGGTGAACTCCACCCGTC	TCGTCGGCGTAGGCATAGAGAT
Gb4CL-F/R	CGACGTCGTAGCGCTTCCTTAT	CTCGTGACCAAACCTTTGTGCG
GbCAD-F/R	CCCGAGAAACTAGCACCAGAGC	GCCGCATTAGAGCTCACAAGGA
GbCHI-F/R	CAATGACTGTGGAGCCGACA	CCCAGAGATGATGATTCAGTGG
GbDFR-F/R	GTTCCCACCAAGTTCAAAGATG	GAAGCAGTCCCTTTTCTCGG
GbTT7-F/R	GCTTGCGAGATCAATGGCTACT	CAGCACCAAATGGAATCAACTC
GhUBQ7-F/R	GAAGGCATTCCACCTGACCAAC	CTTGACCTTCTTCTTCTTGTGCTTG

图1 不同浓度外源褪黑素预处理对棉花枯萎病抗性的影响 A：接种枯萎菌22 d后棉花植株表型；B：接种枯萎菌22 d后棉花剖杆表型；C：接种枯萎菌后不同时间点棉花病情指数。不同字母表示差异显著（P<0.05），下同

Fig. 1 Effects of exogenous melatonin pretreatment at different concentration on the cotton resistance to Fusarium oxysporum f.sp. vasinfectum（Fov） A: Disease phenotypes of cotton with different treatments at 22 d. B: Disease phenotypes in stems of cotton treated with different treatments at 22 d. C: Cotton disease index at different time points of plants treated with different treatments. Different letters indicate significant difference（P <0.05）, the same below

图2 不同浓度外源褪黑素对枯萎菌生长的影响 A：放置7 d后枯萎菌生长情况；B：放置7 d后枯萎菌菌落直径

Fig. 2 Effects of exogenous melatonin at different concentration on the growth of Fov A: Fov colonies 7 d after inoculation on PDA medium. B: Fov colony diameter 7 d after inoculation on PDA medium

图3 褪黑素预处理对接菌后棉花抗氧化系统的影响 * 表示差异显著（P<0.05），** 表示差异极显著（P<0.01），下同

Fig. 3 Effects of melatonin pretreatment on cotton antioxidant system after Fov inoculated * indicates significant difference（P < 0.05）, ** indicates extremely significant difference（P < 0.01）, the same below

图4 褪黑素预处理对接菌后棉花防御酶活性的影响

Fig. 4 Effects of melatonin pretreatment on the cotton defense enzyme activity after Fov inoculated

图5 褪黑素预处理对接菌后棉花木质素、类黄酮代谢通路相关基因表达水平的影响

Fig. 5 Effects of melatonin pretreatment on the cotton expressions of genes related to phenylpropane metabolic pathway related genes after Fov inoculated

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