陆地棉GhCOMT28对干旱胁迫的响应

doi:10.13304/j.nykjdb.2024.0059

中国农业科技导报 ›› 2025, Vol. 27 ›› Issue (4): 45-56.DOI: 10.13304/j.nykjdb.2024.0059

陆地棉GhCOMT28对干旱胁迫的响应

陈宜新¹(), 杨秀波¹, 田士军², 王聪¹, 白志英¹^,³, 李存东¹(), 张科¹()

^1.河北农业大学农学院，省部共建华北作物改良与调控国家重点实验室，河北省作物生长调控实验室，河北保定 071000
^2.河北农业大学实验实训中心，河北保定 071000
^3.河北农业大学生命科学学院，河北保定 071000

收稿日期:2024-01-21 接受日期:2024-03-08 出版日期:2025-04-15 发布日期:2025-04-15
通讯作者: 李存东,张科
作者简介:陈宜新 E-mail：cyx06060701@163.com
基金资助:
河北省自然科学基金项目(C2022204036);河北省省级科技计划资助项目(23567601H);河北省高等学校科学技术研究项目(QN202037);华北作物改良与调控国家重点实验室自主研究课题项目(NCCIR2023ZZ-7);国家自然科学基金项目(31900623)

Response of GhCOMT28 to Drought Stress in Gossypium hirsutum

Yixin CHEN¹(), Xiubo YANG¹, Shijun TIAN², Cong WANG¹, Zhiying BAI¹^,³, Cundong LI¹(), Ke ZHANG¹()

^1.State Key Laboratory of North China Crop Improvement and Regulation，Key Laboratory Crop Growth Regulation of Hebei Province，College of Agronomy，Hebei Agricultural University，Hebei Baoding 071000，China
^2.Experiment and Practice Training center of Hebei Agricultural University，Hebei Agricultural University，Hebei Baoding 071000，China
^3.College of Life Sciences，Hebei Agricultural University，Hebei Baoding 071000 China

Received:2024-01-21 Accepted:2024-03-08 Online:2025-04-15 Published:2025-04-15
Contact: Cundong LI,Ke ZHANG

摘要/Abstract

摘要：

褪黑素作为一种吲哚胺类化合物，对调节植物生长发育及抵御逆境胁迫有广泛作用。COMT（caffeic acid O-methyltransferase）是一种O-甲基转移酶，在苯丙烷代谢途径中发挥重要作用，是褪黑素合成的关键酶。陆地棉中共有57个GhCOMT同源基因，其中GhCOMT28和GhCOMT55与拟南芥AtCOMT同源性最高，以GhCOMT28为例，对其生物学功能进行了解析，发现GhCOMT28和GhCOMT55在棉花根和茎的表达量最高，定位于细胞质。GhCOMT28受干旱胁迫诱导，沉默GhCOMT28（TRV2：GhCOMT28）导致叶片中褪黑素水平降低，超氧化物歧化酶（superoxide dismutase，SOD）、过氧化物酶（peroxidase，POD）、过氧化氢酶（catalase，CAT）等抗氧化酶活性降低，自由基积累增加，植株耐旱性下降。在拟南芥中超表达GhCOMT28（35S：GhCOMT28-GFP）则显著提高植株耐旱性。揭示了GhCOMT28对于棉花幼苗抵御干旱有重要调控作用，在耐旱品种改良中具有利用潜力。

关键词: 棉花, 褪黑素, GhCOMT28, 非生物胁迫, 干旱胁迫

Abstract:

Melatonin， as an indoleamine compound， has a wide range of roles in regulating plant growth and development and resisting adversity stress. COMT （caffeic acid O-methyltransferase） is an O-methyltransferase that plays an important role in the phenylpropane metabolic pathway and is a key enzyme in melatonin synthesis. There are 57 GhCOMTs homologous genes in terrestrial cotton， among which GhCOMT28 and GhCOMT55 have the highest homology with Arabidopsis thalianaAtCOMT. In this study， GhCOMT28 was used as an example to parse its biological function， and GhCOMT28 and GhCOMT55 had the highest expression in the roots and stems of cotton， and GhCOMT28 was localized in the cells in the cytoplasm of cell. GhCOMT28 mRNA was induced by drought stress， and silencing of GhCOMT28 （TRV2：GhCOMT28） resulted in reduced melatonin levels in leaves， and the expression of superoxide dismutase （SOD）， peroxidase （POD）， catalase （CAT） and other antioxidant enzymes decreased， free radicals accumulation increased， and plant drought tolerance decreased. Overexpression of GhCOMT28 （35S：GhCOMT28-GFP） in Arabidopsis significantly improved plant drought tolerance. This study revealed that GhCOMT28 had an important role in regulating drought tolerance in cotton seedlings and had potential use for drought-tolerant variety improvement.

Key words: cotton, melatonin, GhCOMT28, abiotic stress, drought stress

中图分类号:

S562

陈宜新, 杨秀波, 田士军, 王聪, 白志英, 李存东, 张科. 陆地棉GhCOMT28对干旱胁迫的响应[J]. 中国农业科技导报, 2025, 27(4): 45-56.

Yixin CHEN, Xiubo YANG, Shijun TIAN, Cong WANG, Zhiying BAI, Cundong LI, Ke ZHANG. Response of GhCOMT28 to Drought Stress in Gossypium hirsutum[J]. Journal of Agricultural Science and Technology, 2025, 27(4): 45-56.

图/表 10

表1 霍格兰德营养液配方

Table 1 Hoagland nutrient solution formula

药品名称 Name	药品用量 Dosage/（g·L^-1）
Ca（NO₃）₂·4H₂O	118.075
KNO₃	50.550
MgSO₄·7H₂O	49.294
KH₂PO₄	13.609
NH₄H₂PO₄	51.764
H₃BO₃	2.860
MnSO₄·H₂O	1.180
ZnSO₄·7H₂O	0.220
CuSO₄·5H₂O	0.080
（NH₄）₆Mo₇O₂₄·4H₂O	0.020

表2 本研究中所用引物序列

Table 2 Primer sequences used in this study

引物名称 Primer name	引物序列 Primer sequence（5'-3'）	用途 Usage
GhCOMT28-CDS-F	CGGGATCCATGGGTTCAATCGGTGAAACTCA	转基因载体构建 Construct transgenic vector
GhCOMT28-CDS-R	GCTCTAGACAAACACTTTTGAGAAACTCCATG	转基因载体构建 Construct transgenic vector
V-GhCOMT28-F	GGAATTCCGCCTCATGTCATTGAGGATGCT	基因沉默载体构建 Construct gene silence vector
V-GhCOMT28-R	GGAATTCCGTCTGGCAAAGCTTCATAGCAG	基因沉默载体构建 Construct gene silence vector
q-GhCOMT28-F	TTGAGGATGCTCCTAGCTGTCC	qRT-PCR
q-GhCOMT28-R	AATCACTTTCCCGTTGTCTGG
q-GhACTIN7-F	ATCCTCCGTCTTGACCTTG
q-GhACTIN7-R	TGTCCGTCAGGCAACTCAT
q-GhNCED1-F	GCACGACTTCGCCATCACT
q-GhNCED1-R	GGTTCTTCCCAAGCATTCCA
q-GhCBF5-F	ACCACTGCGGTTATGGCTACT
q-GhCBF5-R	CAGTTGGCAACGCGACATTTC
q-GhHSFA2-F	GGGGATTGTAACAACGTCAGC
q-GhHSFA2-R	GCGGTTAATGCTCCAGGAAAC
q-GhNHX1-F	CCAGCATGCTCTCAGACCAA
q-GhNHX1-R	GGAACGAAGGGCACAAAACC

图1 GhCOMT28及其同源蛋白的系统发育分析

Fig.1 Phylogenetic analysis of GhCOMT28 and its homologous protein

图2 GhCOMT28的亚细胞定位注：标尺为50 μm。

Fig.2 Subcellular localization of GhCOMT28Note： The scale is 50 μm.

图3 GhCOMT28的组织表达模式分析

Fig.3 Analysis of GhCOMT28 expression pattern

图4 不同环境胁迫下GhCOMT28的表达模式分析A：干旱（PEG）胁迫；B：冷胁迫；C：高温胁迫；D：盐胁迫。不同小写字母表示不同处理时间基因的表达量之间差异在P<0.05水平显著。

Fig.4 Analysis of expression patterns of GhCOMT28 under different environmental stressA： Drought（PEG） stress； B： Cold stress； C： Heat stress； D： Salt stress. Different lowercase letters indicate that the difference in gene expression between different treatment times is significant at the P<0.05 level.

图5 TRV2：GhCOMT28的耐旱性分析A：TRV2：CLA白化表型；B：GhCOMT28沉默效率检测；C：褪黑素含量检测；D：干旱胁迫7 d后，TRV2：00和TRV2：GhCOMT28存活率统计；E~F：干旱胁迫14 d棉花幼苗的表型； G~H：复水7 d棉花幼苗的表型；*和**分别表示TRV2：GhCOMT28和TRV2：00之间的差异在P<0.05和P<0.01水平显著；标尺为5 cm

Fig. 5 Drought tolerance analysis of TRV2：GhCOMT28A： TRV2： CLA albino phenotype； B： GhCOMT28 silencing efficiency detection； C： Melatonin content detection； D： Survival rate of TRV2： 00 and TRV2： GhCOMT28 after 7 d drought stress； E~F： Phenotype of cotton seedlings after 14 d of drought stress； G~H： Phenotype of cotton seedlings after 7 d of recovery water； * and ** indicate that the difference between the TRV2：GhCOMT28 and TRV2：00 is significant at P<0.05 and P<0.01 levels， respectively； the scale is 5 cm

图6 TRV2：GhCOMT28对叶片抗氧化体系的影响注：CK—正常生长；DS—6% PEG6000模拟干旱胁迫；不同小写字母表示不同样本之间差异在P<0.05水平显著。

Fig.6 TRV2：GhCOMT28 affects the antioxidant system of cotton.Note： CK—Normal growth； DS—6% PEG6000 simulated drought stress； different lowercase letters indicate that the difference between different samples is significant at the P<0.05 level.

图7 棉花叶片的NBT和DAB染色注：CK—正常生长；DS—6% PEG6000模拟干旱胁迫。标尺为2 cm。

Fig. 7 NBT and DAB staining of cotton leavesNote： CK—Normal growth； DS—6% PEG6000 simulated drought stress. Bar is 2 cm.

图8 超表达GhCOMT28增强拟南芥耐旱性A：转基因拟南芥中GhCOMT28-GFP转录水平检测； B：拟南芥植株的存活率统计； C：不同条件下拟南芥的耐旱表型；标尺为5 cm

Fig. 8 Overexpression of GhCOMT28 enhances drought resistance in Arabidopsis thalianaA： Detection of GhCOMT28-GFP transcription level in transgenic Arabidopsis； B： Statistics of survival rate of Arabidopsis plants under drought stress； C：Drought tolerance phenotype of transgenic Arabidopsis under different conditions； the scale is 5 cm

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陆地棉GhCOMT28对干旱胁迫的响应

Response of GhCOMT28 to Drought Stress in Gossypium hirsutum

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