Research Of The Regulation Mechanism Of Exogenous Melatonin On Lignin Deposition In Postharvest Water Bamboo （Zizania Latifolia） During Cold Storage

Water bamboo（Zizania latifolia）is prone to deterioration such as lignification after harvest,which leads to the decline of edible and commercial value,and its storage time is shorten.Lignin deposition is an important cause of lignification.In order to delay the deposition of lignin in water bamboo shoot,maintain its quality and extend the shelf life,this paper used exogenous melatonin to improve the quality of fresh water bamboo during cold storage.By analyzing the changes of storage quality,metabolic pathways of lignin and transcriptome sequencing,the mechanism and related regulatory factors of lignin deposition of water bamboo and the effect of melatonin treatment on this process were investigated.The main findings are as follows:（1）Effects of exogenous melatonin on preservation of water bamboo during cold storageIn this study,100 μM exogenous melatonin was used to treat the post-harvest water bamboo shoots,and the preservation effect was analyzed.Melatonin treatment can delay the browning,hardness and weight loss of water bamboo shoots,maintain the total phenolic content,inhibit lignin deposition and the activities of related enzymes in phenylpropane metabolism such as phenylalanine ammonia lyase（PAL）,cinnamyl alcohol desorption Catalase（CAD）and peroxidase（POD）.The results showed that exogenous melatonin treatment could significantly improve the quality of water bamboo during cold storage.（2）Effects of exogenous melatonin on transcription level of water bamboo during cold storage and the identification of regulatory factors of lignin depositionHigh-throughput sequencing technology was used to sequence the transcriptome of 0,14,28 d control and 14 d melatonin-treated water bamboo during the cold storage,and perform functional annotation and enrichment analysis of differentially expressed genes.It was found that the deposition of lignin depended on lignin biosynthesis,and multiple metabolic pathways were involved,the most important of which was the phenylpropane pathway（ko00940）.By analyzing the expression levels of genes of important enzymes in the phenylpropane metabolic pathway,it was found that ZIPAL,ZlCAD3,ZlPOD2,and ZlPOD3 showed an upward trend,which was consistent with the changes in the corresponding enzyme activities and with the change trend of lignin accumulation during storage.The expression of nine respiratory burst oxidase homologs（RBOH）genes in the RBOHs gene family of water bamboo was increased during postharvest cold storage,suggesting that RBOH-mediated ROS generation was the main reason for the lignification.Melatonin treatment can effectively inhibit the lignin deposition of water bamboo during cold storage by down-regulating lignin deposition-related genes and reducing the expression of ROS-related RBOHs genes,thereby delaying the process of lignification during cold storage.（3）Effects of RBOH on phenylpropane metabolism during lignification of water bamboo during cold storageIn this study,diphenyl iodide chloride（DPI）and hydrogen peroxide（H2O2）were used to treat the water bamboo,and it was found that DPI treatment could inhibit the expression of phenylpropane metabolism-related genes and RBOHs gene family in water bamboo during cold storage,while H2O2 treatment could improve its relative expression and accelerate the lignification process.During the cold storage,the lignin staining area and lignin content had similar trends,which indicated that the lignin was mainly deposited in its lower cortex.The lignin staining area and lignin content of DPI treatment were significantly smaller and lower than those in the control group,while H2O2 treatment increased the lignin staining area and content.The accumulation of ROS is closely related to the lignification in water bamboo,which indicates the regulatory function of the RBOHs gene family on lignin biosynthesis.（4）Verification of ZIRBOHA1 gene function in the lignin deposition processIn this study,ZlRBOHA1 was selected from the RBOHs gene family for functional verification.The overexpression vector pCAMBIA1303-ZlRBOHA1 was constructed to characterize Arabidopsis thaliana,and it was found that ZlRBOHA1 overexpression promoted Arabidopsis thaliana lignin biosynthesis.