过表达SlWRKY41提高番茄幼苗抗旱性

doi:10.13560/j.cnki.biotech.bull.1985.2024-0688

摘要/Abstract

摘要：

目的探究SlWRKY41转录因子在番茄响应干旱胁迫中的作用，为进一步丰富番茄抗逆种质资源和选育抗旱品种提供理论依据。方法以野生型番茄‘Ailsa Craig’为材料，以其cDNA为模板克隆SlWRKY41基因，构建OE-SlWRKY41转基因番茄株系，对野生型和OE-SlWRKY41转基因番茄株系进行干旱处理，在处理第4、8天时对其进行表型观察和相关生理、生化指标的测定，并进行胁迫相关基因的表达分析。结果与野生型植株相比，干旱胁迫8 d后，OE-SlWRKY41转基因植株的叶片卷曲，萎蔫程度较轻，膜脂过氧化水平显著降低；气孔导度和蒸腾速率显著降低，净光合速率显著提高；根系形态指标整体呈现上升趋势，叶片相对含水量、抗氧化酶活性和渗透调节物质（可溶性糖和脯氨酸含量）显著增加，相对电导率显著降低；过表达SlWRKY41植株中的胁迫响应基因（SlSOD、SlPOD和SlP5CS1）显著上调。结论 SlWRKY41能够响应干旱，过表达SlWRKY41能够提高番茄幼苗对干旱胁迫的耐受性。

关键词: 番茄, 转录因子, SlWRKY41, 干旱胁迫, 光合, 抗氧化系统, 渗透调节

Abstract:

Objective To explore the role of SlWRKY41 transcription factor in response to drought stress in tomato, and to provide theoretical value and important basis for the enrichment of tomato germplasm resources and the selection and breeding of drought-resistant varieties of tomato. Method The experiment was conducted with wild-type tomato 'Ailsa Craig' as the material, and its cDNA was used as the template to clone the SlWRKY41 gene to construct the OE-SlWRKY41 transgenic tomato lines, and the wild-type and OE-SlWRKY41 transgenic tomato lines were subjected to drought stress treatments, and the phenotypic observations and related physiological and biochemical indexes were determined at the day 4 and 8 of the treatment. Result Compared with wild-type plants, after 8 d of drought stress, OE-SlWRKY41 transgenic plants had curled leaves with less wilting and significantly lower levels of membrane lipid peroxidation. Stomatal conductance and transpiration rates were significantly lower, and the net photosynthetic rate was significantly higher. The root morphology indexes showed an overall upward trend, and the leaf relative water content, antioxidant enzyme activities and osmotic regulating substances (soluble sugars and proline content) increased significantly, and relative conductivity decreased significantly. The stress response genes (SlSOD, SlPOD and SlP5CS1) in the plants overexpressing SlWRKY41 were significantly up-regulated. Conclusion SlWRKY41 respond to drought, and the overexpression of SlWRKY41 can improve the tolerance of tomato seedlings to drought stress.

Key words: tomato, transcription factors, SlWRKY41, drought stress, photosynthesis, antioxidant system, osmoregulation

刘洁, 王飞, 陶婷, 张玉静, 陈浩婷, 张瑞星, 石玉, 张毅. 过表达SlWRKY41提高番茄幼苗抗旱性[J]. 生物技术通报, 2025, 41(2): 107-118.

LIU Jie, WANG Fei, TAO Ting, ZHANG Yu-jing, CHEN Hao-ting, ZHANG Rui-xing, SHI Yu, ZHANG Yi. Overexpression of SlWRKY41 Improves the Tolerance of Tomato Seedlings to Drought[J]. Biotechnology Bulletin, 2025, 41(2): 107-118.

图/表 10

表1 RT-qPCR引物序列

Table 1 Primer sequences for RT-qPCR

引物名称Primer name	引物序列Primer sequence（5′-3′）
SlActin-F	ACCACTGAGCACAATGTTACCG
SlActin-R	GTCCTCTTCCAGCCATCCA
SlWRKY41-F	GGCTTGAAGGTCCATGTGAAGATG
SlWRKY41-R	ACCCCTGTGTGTTCCTATGAGTAC
SlSOD-F	AATTCATCATTGTGGCAGCA
SlSOD-R	GCCCTTAAGGACAGCAACAG
SlPOD-F	GGTCCAACATGGCAAGTTCT
SlPOD-R	ACATCTTGCCCTTCCAAATG
SlP5CS1-F	TGATGGAAGATTAGCACTTGGAA
SlP5CS1-R	CAACACCTACAGCACCAGAA

图1 SlWRKY41转基因番茄的遗传转化和转基因番茄PCR和RT-qPCR检测A：制备外植体；B：分化愈伤组织；C：生成抗性芽；D：生根培养；E：T1代SlWRKY41转基因番茄基因组DNA PCR检测。M：DNA maker DL2000；1：阴性对照；2：野生型植株；3-8：SlWRKY41转基因株系；9：质粒对照；F：RT-qPCR 检测T2代转基因番茄植株中WRKY41的表达量，不同小写字母代表不同处理间差异达到显著水平（P<0.05），下同

Fig. 1 Genetic transformation of SlWRKY41-transgenic tomato plants and PCR and RT-qPCR detection of transgenic tomato plantsA: Preparation of exosomes. B: Differentiation of healing tissues. C: Generation of resistant buds. D: Rooting culture. E: SlWRKY41 transgenic tomato genomic DNA PCR assay for the T1 generation. M: DNA maker DL2000; 1: negative control; 2: wild-type plants; 3-8: SlWRKY41 transgenic lines; 9: plasmid control. F: RT-qPCR detection of WRKY41 expression in T2 generation transgenic tomato plants, and the different lowercase letters indicate significant levels of difference among treatments(P<0.05), the same below

图2 干旱胁迫后野生型植株和过表达SlWRKY41植株的表型观察OE-WRKY41从左往右依次为OE-WRKY41-1、OE-WRKY41-2

Fig. 2 Phenotypic observations of wild-type and SlWRKY41-overexpressed plants under drought stressOE-WRKY41 from left to right is OE-WRKY41-1，and OE-1WRKY41-2

表2 干旱胁迫对过表达SlWRKY41番茄幼苗生物量的影响

Table 2 Effects of drought stress on the biomasses of tomato seedlings overexpressing SlWRKY41

处理

Treatment

株高

Plant height/cm

茎粗

Stem diameter/mm

地上鲜重Shoot fresh weight/g

地下鲜重 Root fresh weight/g

地上干重Shoot dry weight/g

地下干重 Root dry weight/g

总干重 Total dry weight/g

总鲜重 Total fresh weight/g

表3 干旱胁迫对过表达SlWRKY41番茄幼苗根系形态指标的影响

Table 3 Effects of drought stress on the root system indexes of tomato seedlings overexpressing SlWRKY41

处理

Treatment

根长

Root length/cm

根系表面积

Root surface area/cm²

根平均直径

Mean root diameter/mm

根系体积

Root volume/ cm³

根尖数

Root tips

根分叉数

Root forks

交叉数

Root crosses

图3 干旱胁迫对过表达SlWRKY41番茄幼苗叶片光合指标的影响

Fig. 3 Effects of drought stress on photosynthetic indexes in the leaves of tomato seedlings overexpressing SlWRKY41

图4 干旱胁迫对过表达SlWRKY41番茄幼苗叶片相对电导率和相对含水量的影响

Fig. 4 Effects of drought stress on the relative electrical conductivity and relative water content in the leaves of tomato seedlings overexpressing SlWRKY41

图5 干旱胁迫对过表达SlWRKY41番茄幼苗叶片丙二醛、活性氧含量及抗氧化酶活的影响

Fig. 5 Effects of drought stress on malondialdehyde， reactive oxygen species content and antioxidant enzyme activities in the leaves of tomato seedlings overexpressing SlWRKY41

图6 干旱胁迫对过表达SlWRKY41番茄幼苗叶片可溶性糖和脯氨酸含量的影响

Fig. 6 Effects of drought stress on soluble sugar content and proline in the leaves of tomato seedlings overexpressing SlWRKY41

图7 干旱胁迫下过表达SlWRKY41番茄幼苗胁迫相关基因的相对表达量变化

Fig. 7 Relative expression variations of stress-related genes in the tomato seedlings overexpressing SlWRKY41 under drought stress

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