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Article

Selenium Treatment Regulated the Accumulation of Reactive Oxygen Species and the Expressions of Related Genes in Postharvest Broccoli

by
Yaping Liu
1,2,*,
Wei Wang
1,2,
Gang Ren
1,2,
Yanan Cao
1,2,
Jianbing Di
1,2,
Yu Wang
1,2 and
Lixin Zhang
1,2
1
College of Food Science and Engineering, Shanxi Agricultural University, Jinzhong 030800, China
2
Shanxi Center of Technology Innovation for Storage and Processing of Fruit and Vegetable, Jinzhong 030800, China
*
Author to whom correspondence should be addressed.
Agronomy 2024, 14(5), 1047; https://doi.org/10.3390/agronomy14051047
Submission received: 27 April 2024 / Revised: 10 May 2024 / Accepted: 13 May 2024 / Published: 14 May 2024
(This article belongs to the Special Issue Molecular Regulation Mechanism of Ripening, Senescence and Stress Resistance in Fruits and Vegetables)
Browse Figures
Versions Notes

Abstract

:
This study aimed to investigate the impact of selenium (Se) treatment on the accumulation of reactive oxygen species (ROS) and the expressions of related genes in broccoli. To achieve this, one group of broccoli heads was treated with a selenite solution of 2 mg L−1, while another group was soaked in distilled water, serving as the control. The effects of these treatments were evaluated by analyzing the browning, hydrogen peroxide (H2O2) and malondialdehyde (MDA) contents, enzyme activity, and gene expression levels of WARK and RBOH. Our results show that the Se treatment effectively inhibited H2O2 accumulation in the broccoli and reduced harmful MDA levels. The inhibition of ROS accumulation following the Se treatment was associated with enhanced activity of the CAT and SOD enzymes, increased expression levels of BoCAT and BoSOD, and decreased expression levels of the WRKY and RBOH transcription factors. Our study provides insights into the mechanism of action of selenium and its potential application in vegetable storage.

1. Introduction

Broccoli (Brassica oleracea L. var. Italica) is an important global vegetable, with extensive cultivation covering approximately 140,000 hectares in China alone [1]. Its popularity stems from being a rich source of essential minerals, amino acids, dietary fiber, and various phytochemicals, making it a highly nutritious choice [2]. However, postharvest broccoli faces a challenge due to its susceptibility to oxidative damage, leading to browning at the incision site and significantly reducing its shelf-life. The degree of browning in cut broccoli directly impacts its commercial value as it affects consumers’ willingness to purchase.
ROS are natural byproducts of plant metabolism, serving as both signaling molecules and potential toxins [3]. Under chilling stress, plants experience excessive ROS accumulation in cells, including superoxide anion (O2−) and hydrogen peroxide (H2O2) [4]. However, elevated ROS levels can lead to oxidative damage to macromolecules and contribute to cell death pathways [5]. Generally, plants possess intricate mechanisms to maintain a delicate balance of ROS as they can have both beneficial and harmful effects. Enzymes such as catalase (CAT), ascorbate peroxidase (APX), and superoxide dismutase (SOD) function in living organisms to regulate ROS concentrations by scavenging H2O2 and O2− [6,7]. Among the various pathways of ROS production, the NADPH oxidase encoded by the respiratory burst oxidase homolog (RBOH) gene has been extensively studied [8]. Under abiotic stress conditions, NADPH oxidase is activated by calcium and phosphorylation, leading to ROS production in plants [9]. The release of ROS is closely associated with the activity of specific cis-acting elements in the upstream regions of target genes. Among those elements, the W-Box has been identified as a main ROS-specific element [10]. Interestingly, the W-Box (TTGACC) has been characterized in several members of the ROBHs gene family [11]. Previous research by [12] demonstrated that the mutation of AtRBOHd in Arabidopsis limited ROS accumulation, while [13] showed that the accumulation of RBOHd protein increased through the mutation of PIRE and PBL13, leading to enhanced ROS production. Additionally, WRKY TFs have been identified as positive regulators of ROBH genes. For instance, Adachi et al. [14] reported that the silencing of WRKYs resulted in the downregulation of ROBHb, impairing ROS burst, with the W-Box identified as a binding site for WRKYs in the cis-acting elements of ROBH genes.
Selenium is widely recognized for its potential antioxidant properties and has been extensively studied [15]. Studies investigating selenium’s antioxidant effects date back to the 1970s and 1980s, during which it was proposed that selenium occupies active sites in glutathione peroxidase [16]. Subsequent research has established a consensus that selenium, as a component of various selenoproteins with antioxidative activity, plays a crucial role in the response to oxidative stress [17]. These selenoproteins include glutathione peroxidase and thioredoxin reductase [18]. Of particular importance, selenium has been reported to enhance stress resistance, antioxidant capacity, and enzyme activity in plants [19]. Despite these promising findings, there is a limited information on the application of selenium in postharvest fruit and vegetable preservation.
In our previous study, we employed transcriptome sequencing (RNA-seq) and observed a significant upregulation of genes encoding BoPOD, BoSOD, BoAPX, and BoCAT after selenium treatment. In contrast, the expressions of BoRBOHd and BoRBOHf, which are closely associated with ROS concentration, were downregulated. These findings led us to speculate that selenium may regulate ROS levels by modulating the expressions of ROS metabolism-related genes in broccoli. Therefore, the main objective of this study was to investigate the impact of selenium on antioxidation in broccoli. Through our investigation, we aimed to gain deeper insights into the potential mechanisms underlying selenium’s biological functions in controlling antioxidation in broccoli.

2. Materials and Methods

2.1. Plant Material, Processing, and Storage Conditions

For this experiment, we collected fresh broccoli samples from the Juxin Entrepreneurship Park of Shanxi Agricultural University, ensuring that they were free from any surface bruises, pest infestation, or mechanical damage. The selected broccoli heads were washed with distilled water and left to naturally air dry. Subsequently, the broccoli heads were randomly divided into two groups, with each group containing 35 broccoli heads. One group was immersed in water containing 2 mg L−1 of selenite (Se treatment, Damao chemical reagent factory, Tianjin, China), while the other group was immersed in distilled water (control) for 20 min. Following the dipping process, the broccoli heads were air-dried and carefully placed in polyethylene film bags (0.02 mm thickness, 80 cm × 60 cm in size) before being stored at 0 °C.
During the storage period, flower buds were excised from the broccoli at different time intervals, immediately frozen in liquid nitrogen, and stored at −80 °C until further analysis. Sampling of the flower buds was conducted randomly from five broccoli samples every 10 days, up to a storage duration of 60 days. For each parameter analysis, three biological replicates were conducted.

2.2. Malondialdehyde (MDA) and H2O2 Content Analysis

The measurement of the MDA content was conducted in accordance with the thiobarbituric acid (TBA) method [20]. One gram of flower buds from five broccoli heads was utilized. The MDA content was expressed as μ mol kg−1 on a fresh weight (FW) basis.
To determine the H2O2 content, 0.1 g of broccoli head was employed, and the Hydrogen Peroxide Assay Kit (Solarbio, Beijing, China) was used. The H2O2 content was subsequently quantified spectrophotometrically using an H2O2-specific reagent, titanium sulfate.

2.3. Enzyme Activities Assays

The activities of the POD, CAT, SOD, and APX enzymes were measured using an enzyme activity assay kit (Solarbio, Beijing, China), following the manufacturer’s instructions. We strictly adhered to all steps and procedures outlined in the provided instructions to ensure accurate and reliable enzyme activity measurements.

2.4. RNA-seq Analysis

The broccoli samples were analyzed by RNA-seq following the established research methods as described in [19].

2.5. RT-qPCR Analysis

The total RNA was extracted from the broccoli florets using the TransZol Up Plus RNA Kit (TransGen, Beijing, China). Gene-specific primers, as listed in the Supplementary Materials Table S1, were employed for the qRT-PCR analysis, and the qRT-PCR reactions were performed using the PerfectStart Green qPCR SuperMix (TransGen, Beijing, China). The gene-specific primer sequences for qRT-PCR were provided in Table S1. To ensure accurate normalization, the tomato Actin gene (LOC) was used as the internal control. The relative gene expression values were calculated using the 2−ΔΔCt method, and the results were normalized using the Actin internal reference gene. The analysis was conducted with three independent biological replicates.

2.6. Evolutionary Tree Construction of BoRBOH and BoWRKY

The standalone BLAST package (NCBI, www.ncbi.nlm.nih.gov/blast/, accessed on 12 March 2023) was used for the homolog search analysis of the BoRBOH and BoWRKY genes. Phylogenetic trees were constructed using the neighbor-joining (NJ) method with 1000 bootstrap replicates in MEGA 7.0 software (Mega Limited, Auckland, New Zealand).

2.7. Statistical Analysis

Statistical and linear regression analyses were performed using Origin Pro 2021 (Origin Lab Inc., Northampton, MA, USA). Standard deviation calculations, one-way ANOVA, and Pearson correlation analysis were performed using SPSS v.20 software (SPSS Inc., Chicago, IL, USA).

3. Results

3.1. Browning, H2O2 Content, and MDA Content

As shown in Figure 1A, the browning of broccoli increased during storage. However, the Se treatment significantly reduced the browning compared to the control group throughout the storage period. The H2O2 content increased during the first twenty days of storage in the control group, followed by a decrease after twenty days (Figure 1B). A similar trend was observed in the Se-treated broccoli head group. However, from day 40 to 60, the Se treatment significantly inhibited the accumulation of H2O2 compared to the control group. Additionally, the H2O2 content in the Se-treated broccoli head was higher than in the control broccoli head at 20 d.
The MDA content in the broccoli head of the control group reached its highest peak at 20 d, after which it decreased gradually (Figure 1C). While the MDA content in the Se treatment group showed a similar trend to the control, it was consistently significantly lower than that in the control broccoli head throughout the entire storage period.

3.2. POD, CAT, SOD, and APX Activity

In general, the POD activity of Se−treated broccoli head was lower than that of the control group during the first thirty days of storage (Figure 2A). Conversely, the Se treatment resulted in higher CAT activity and SOD activity than observed in the control group throughout the entire storage period (Figure 2B,C). However, the APX activity in the control group increased during storage, while the Se treatment resulted in lower APX activity compared to the control group during most of the storage period (except at 50 days) (Figure 2D).

3.3. RNA-seq Results Analysis

The RNA-seq results show that there were two and five differentially expressed genes related to ROS metabolism for the plant RBOH gene and WRKY gene families, respectively. These genes participate in multiple biological processes, including the MAPK signaling pathway and plant–pathogen interaction (Figure S1).

3.4. Genes Expression Level

As shown in Figure 3A, the BoPOD gene expression in the Se-treated broccoli head was generally lower than in the control during storage, except at 30 d and 40 d. The expression of BoPOD rapidly peaked at 40 d in the Se treatment, while it appeared at 50 d in the control. In the control broccoli heads, the expression of BoSOD rapidly increased within the first ten days of storage, followed by a decrease, maintaining a lower level. In contrast, the expression of BoSOD in the Se-treated broccoli head remained relatively stable level and was higher compared to the control group from 20 d to 40 d of storage, although its expression level was lower than the control at 10 d (Figure 3B). Similarly, the BoAPX expression in the control group rapidly increased at 10 d and then gradually decreased. Conversely, in the Se-treated group, the expression of BoAPX showed a rapid decrease at 10 d. Unlike BoSOD, the BoAPX expression in the Se-treated broccoli heads was higher than in the control from 40 d to 60 d of storage (Figure 3C). Moreover, the BoCAT expression in the Se-treated broccoli heads was higher than that in the control broccoli heads for most of the storage time (Figure 3D).

3.5. Expression Patterns of BoRBOHd and BoRBOHf

Based on similarity searches using broccoli’s RBOHd and RBOHf amino acid sequences, two homologous RBOH genes were identified from Arabidopsis (Figure S2). From a general perspective, the expression patterns of RBOH-related genes were found to be similar. Specifically, for BoRBOHf, its expression level in the Se-treated broccoli heads was lower than that in the control broccoli heads after 20 days of storage (Figure 4A). Additionally, the expression of BoRBOHf was induced at low temperatures, particularly at 0 and 20 days of storage. On the other hand, the expression of BoRBOHd showed a durable induction, especially at 0 and 40 days of storage (Figure 4B). Overall, the data in Figure 4 demonstrate that the Se treatment effectively inhibited the expressions of both BoRBOHf and BoRBOHd during cold storage.

3.6. Expression Patterns of BoWRKYs

The homology analysis results revealed a striking similarity between BoWRKY33, BoWRKY25, BoWRKY15, and BoWRKY6, and their respective homologs in Arabidopsis, namely, AtWRKY33, AtWRKY25, AtWRKY15, and AtWRKY6 (Figure S3). Throughout the storage period, except BoWRKY6, the expression levels of all WRKY-related genes exhibited an initial increase, followed by a subsequent decrease. Notably, the expression of BoWRKY33 was significantly inhibited upon treatment with Se during storage (Figure 5A). Additionally, the expressions of BoWRKY25 and BoWRKY15 were inhibited by the Se treatment at 0 and 40 days of storage, respectively (Figure 5C,E). Interestingly, BoWRKY6 expression showed an upward trend, and there were significant differences in its expression level between the Se treatment and the control groups (Figure 5B). Importantly, the qPCR results demonstrate a high level of concordance with the RNA-Seq data, underscoring the reliability of our RNA-Seq findings.

4. Discussion

In recent years, cruciferous vegetables have gained significant attention for their ability to accumulate Se, making them stand out among other vegetation in this regard [21]. Studies have demonstrated that cruciferous vegetables can accumulate Se at a rate 100 times higher than other plants in the vicinity [22]. Selenium is a beneficial element known to enhance plant growth and development by inducing secondary metabolism and improving the antioxidant capacity of enzymes [23]. While there are numerous studies focusing on the effects of Se biofortification on crop growth and development, there is limited research on the impact of postharvest Se treatment on the storage quality of broccoli. In our previous studies, we discovered that Se treatment not only preserved the green phenotype of broccoli heads but also inhibited the loss of aromatic compounds during storage, resulting in improved storage quality [19,24]. However, a comprehensive understanding of the storage quality effects of Se treatment requires further exploration. Therefore, in the present study, we examined the impact of Se treatment on the accumulation of ROS in broccoli heads during storage. The results revealed that Se application at concentrations of 2 mg L−1 enhanced the total antioxidant capacity of the broccoli heads, reduced ROS accumulation, and effectively alleviated the browning of wounds in the broccoli heads (Figure 1). To gain insights into the metabolic mechanisms underlying ROS regulation under Se treatment, we conducted transcription analysis. Our findings indicate that the expressions of genes belonging to the BoWRKY and BoRBOH transcription factor families were influenced by the Se treatment, thereby affecting the concentration of ROS in broccoli heads.
The cellular presence of ROS, such as H2O2, O2-, singlet oxygen (O2), hydroxyl radical (HO), and various forms of organic and inorganic peroxides [25], can lead to damage in lipids, proteins, RNA, DNA, and other cellular molecules. One major form of damage is membrane lipid peroxidation, leading to the accumulation of MDA [26]. However, a dynamic equilibrium exists between ROS and the antioxidant defense system in plants. Within this equilibrium, to mitigate the harmful effects of MDA, the activity of antioxidant enzymes, such as CAT, SOD, and PPO, is modulated [27]. In this study, postharvest selenium treatment was found to significantly reduce the H2O2 and MDA contents in the broccoli heads compared to the control group (Figure 1B,C). Meanwhile, we observed increases in the activities of the CAT, SOD, and POD enzymes following the Se treatment, which facilitated the breakdown of H2O2 and O2− (Figure 2A,C). Additionally, the metabolism of ROS was closely linked to the process of browning, and increasing the antioxidant capacity has been shown to delay browning [28]. This observation was supported by a study on sweet potatoes, which revealed the accumulation of a significant amount of H2O2 in cells from browning regions [29], suggesting a direct association between ROS and browning. Our experimental results further substantiate these findings, demonstrating a positive correlation between browning and H2O2 content, as well as MDA content and H2O2 content. Moreover, our study highlights that the postharvest Se treatment effectively delayed browning by scavenging H2O2, thus contributing to improved storage quality in broccoli heads.
Extensive insights into the metabolic pathway (KEGG) were obtained from the analysis of the transcriptomic data, revealing the involvement of pathways related to ROS metabolism, including the plant–pathogen Interaction and MAPK signaling pathways. The critical sources of ROS in plants are NADPH oxidases (RBOHs) [30]. These transmembrane proteins play a highly regulated role, utilizing cytosolic NADPH to generate O2− in the apoplast, which is subsequently converted to H2O2 [6]. Emerging evidence indicates that RBOH responds to various biotic and abiotic stresses primarily by modulating ROS generation [31]. For example, in tomato, exposure to high-temperature and oxidative stress leads to an increase in the expression level of SIRBOHf, resulting in the accumulation of ROS [32]. Similarly, low-oxygen and low-temperature stresses trigger an upregulation of RBOHd [33]. In this study, we identified two BoRBOHs, BoRBOHd and BoRBOHf, through our transcriptomics analysis of broccoli. Subsequent RT-qPCR analysis demonstrated that BoRBOHd and BoRBOHf were significantly induced by low temperature but inhibited by the Se treatment during storage. Although low-temperature storage is a common preservation method, it renders fruits and vegetables susceptible to cold stress during storage. Furthermore, we observed high homology between BoRBOHd and AtRBOHd, as well as between BoRBOHf and AtRBOHf, suggesting similar functions between these pairs. It is noteworthy that AtRBOHd and AtRBOHf have been verified as primary contributors to ROS generation under abiotic stresses [34]. In summary, the application of Se has proven effective in alleviating cold stress in broccoli by inhibiting the overexpression of BoRBOHd and BoRBOHf during low-temperature storage.
WRKY genes play a crucial and widespread role in higher plants, influencing various aspects of plant growth and development, metabolic processes, morphological and structural formation, and signal transduction [35]. These transcription factors are an integral part of the plant’s response to environmental stresses, regulating the expressions of hormone-associated genes either positively or negatively, and reacting to signals from plant pathogen infestation [36]. In addition, WRKY genes are implicated in the plant’s response to abiotic stresses. For instance, in Arabidopsis, AtWRKY25, AtWRKY26, and AtWRKY33 are induced under cold stress [37]. Furthermore, AtWRKY25 and AtWRKY33 have been identified as substrates of the protein kinase MPK4, which stimulates H2O2 production [38]. By conducting a sequence alignment of BoWRKY and their homologs, we observed a high degree of similarity between BoWRKY33 and AtWRKY33, as well as between BoWRKY25-1, BoWRKY25-2 and AtWRKY25. Additionally, our investigation revealed that BoWRKY33 and BoWRKY25 are induced under low-temperature conditions. Surprisingly, their expression levels decreased following the Se treatment. Furthermore, existing studies have indicated that the expression of RBOHd is regulated by WRKY33, which consequently affects ROS production indirectly [14]. Similarly, AtWRKY6 and AtWRKY15, which share high homology with BoWRKY6 and BoWRKY15, respectively, have been reported to be induced by ROS. Decreasing the expression of AtWRKY15 was found to enhance resistance to oxidative and osmotic stress [39]. In our experiments, we observed that the expression levels of BoWRKY6 and BoWRKY15 were lower compared to the control group. This could potentially be attributed to the relatively low concentration of ROS in the samples. Overall, the Se treatment appears to effectively inhibit the accumulation of ROS by modulating the expressions of the BoWRKY33 and BoWRKY25 transcription factors.

5. Conclusions

Our experimental findings demonstrate that Se treatment effectively mitigated ROS accumulation in postharvest broccoli during cold storage. Compared to the control group, the Se-treated broccoli exhibited lower levels of excessive ROS accumulation and membrane lipid peroxidation. Notably, the expression levels of BoRBOHd and BoRBOHf, which are known to be associated with ROS production, were reduced in response to the Se treatment. Additionally, the Se treatment seemed to regulate the activity of the BoWRKY33 and BoWRKY25 transcription factors, contributing to the inhibition of ROS production. We speculate that the expression of BoRBOH might be affected by the levels of the BoWRKY transcription factors, warranting further investigation. Our findings provide novel insights into the regulatory mechanisms of Se treatment in scavenging ROS in postharvest broccoli at the transcriptional level. Understanding these molecular interactions and signaling pathways could lead to the development of strategies to improve the postharvest storage quality and stress tolerance of broccoli and other fruits or vegetables.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/agronomy14051047/s1, Table S1. Name and sequence of primers designed for genes related to ROS metabolism. Figure S1. Plant-Pathogen interaction and MAPK signaling pathways. Figure S2. Phylogenetic relationships among RBOHs. Phylogenetic tree of RBOH sequences of broccoli (BoRBOH), Arabidopsis thaliana (AtRBOH), Solanum tuberosum (StRBOH), and Oryza sativa (OsRBOH) based on a conservative approximate alignment method. Figure S3. Phylogenetic relationships among WRKYs. Phylogenetic tree of WRKY sequences of broccoli (BoWRKY), Arabidopsis thaliana (AtWRKY), Hordeum vulgare (HvWRKY), zea mays (ZmWRKY) and Oryza sativa (OsWRKY) based on a conservative approximate alignment.

Author Contributions

Conceptualization, W.W., G.R. and Y.L.; methodology, W.W. and G.R.; software, W.W., G.R. and Y.L.; validation, W.W., Y.L. and Y.C.; formal analysis, Y.L. and J.D.; writing—original draft preparation, W.W.; writing—review and editing, Y.L., Y.W. and G.R.; project administration, Y.L. and L.Z. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by the Key Research and Development Plant (Agricultural Field) of Shanxi Province (no. 202102140601017).

Data Availability Statement

The data presented are contained within the article.

Acknowledgments

Thanks to Yan Kexing and Dong Shuai for their help with the molecular experiments.

Conflicts of Interest

The authors declare no conflict of interest.

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Agronomy 14 01047 g001
Figure 1. Effects of Se treatment on browning (A), H2O2 content (B), and MDA content (C) of broccoli during storage. Each data point represents the mean of three replicate assays ± standard error. Different letters represent significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
Figure 1. Effects of Se treatment on browning (A), H2O2 content (B), and MDA content (C) of broccoli during storage. Each data point represents the mean of three replicate assays ± standard error. Different letters represent significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
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Figure 2. Effects of Se treatment on the enzymatic activity of POD (A), CAT (B), SOD (C), and APX (D) in broccoli during storage. Each data point represents the mean of three replicate assays ± standard error. Different letters indicate significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
Figure 2. Effects of Se treatment on the enzymatic activity of POD (A), CAT (B), SOD (C), and APX (D) in broccoli during storage. Each data point represents the mean of three replicate assays ± standard error. Different letters indicate significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
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Figure 3. Effects of Se treatment on the expression levels of genes involved in ROS metabolism, including BoPOD (A), BoSOD (B), BoAPX (C) and BoCAT (D). Each data point represents the mean of three replicate assays ± standard error. Different letters represent significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
Figure 3. Effects of Se treatment on the expression levels of genes involved in ROS metabolism, including BoPOD (A), BoSOD (B), BoAPX (C) and BoCAT (D). Each data point represents the mean of three replicate assays ± standard error. Different letters represent significant differences within the same group during storage (p < 0.05), as determined by Duncan’s multiple range test.
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Figure 4. Effects of Se treatment on the expression levels of RBOH transcription factors involved in ROS metabolism, including BoRBOHf (A) and BoRBOHd (B). Each data point represents a mean of three replicate assays ± standard error. Different letters represent significant differences in the same group during storage (p < 0.05) by Duncan’s multiple range test.
Figure 4. Effects of Se treatment on the expression levels of RBOH transcription factors involved in ROS metabolism, including BoRBOHf (A) and BoRBOHd (B). Each data point represents a mean of three replicate assays ± standard error. Different letters represent significant differences in the same group during storage (p < 0.05) by Duncan’s multiple range test.
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Figure 5. Effects of Se treatment on the expression levels of WRKY transcription factors involved in ROS metabolism, including BoWRKY33 (A), BoWRKY6 (B), BoWRKY25-1 (C), Bo25-2 (D) and BoWRKY15 (E). Each data point represents a mean of three replicate assays ± standard error. Different letters represent significant differences in the same group during storage (p < 0.05) by Duncan’s multiple range test.
Figure 5. Effects of Se treatment on the expression levels of WRKY transcription factors involved in ROS metabolism, including BoWRKY33 (A), BoWRKY6 (B), BoWRKY25-1 (C), Bo25-2 (D) and BoWRKY15 (E). Each data point represents a mean of three replicate assays ± standard error. Different letters represent significant differences in the same group during storage (p < 0.05) by Duncan’s multiple range test.
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MDPI and ACS Style

Liu, Y.; Wang, W.; Ren, G.; Cao, Y.; Di, J.; Wang, Y.; Zhang, L. Selenium Treatment Regulated the Accumulation of Reactive Oxygen Species and the Expressions of Related Genes in Postharvest Broccoli. Agronomy 2024, 14, 1047. https://doi.org/10.3390/agronomy14051047

AMA Style

Liu Y, Wang W, Ren G, Cao Y, Di J, Wang Y, Zhang L. Selenium Treatment Regulated the Accumulation of Reactive Oxygen Species and the Expressions of Related Genes in Postharvest Broccoli. Agronomy. 2024; 14(5):1047. https://doi.org/10.3390/agronomy14051047

Chicago/Turabian Style

Liu, Yaping, Wei Wang, Gang Ren, Yanan Cao, Jianbing Di, Yu Wang, and Lixin Zhang. 2024. "Selenium Treatment Regulated the Accumulation of Reactive Oxygen Species and the Expressions of Related Genes in Postharvest Broccoli" Agronomy 14, no. 5: 1047. https://doi.org/10.3390/agronomy14051047

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