Transcriptomic Analysis Of Exogenous NAA Application To Regulate Graft Healing In Oriental Melon

Oriental melon is an important horticultural crop with high economic value.However,in the actual production of melon,problems such as continuous cropping obstacles and soil-borne diseases have caused decreased yield and reduced quality.Grafting is one of the most effective methods for controlling major diseases.However,there are also phenomena such as increased cost and long incubation period in the cultivation of melon grafted seedlings.Studies have shown that the application of exogenous plant hormones can effectively promote the graft healing process,short the time of grafted healing,and improve the quality of grafted seedlings.Therefore,in this experiment,the grafted seedlings that graft union dipped 40 mg?L-1NAA were used as the treatment,and those dipped water were used as the control.We used the oriental melon 'Yinquan No.1' as the scion,and the white-seed pumpkin 'Shengzhen No.1' as the rootstock.Splice approach was used as the grafting method.Firstly,the paraffin section technique was used to screen three critical stages of melon graft healing process(isolated layer formation,callus formation,vascular bundle formation).Secondly,real-time quantitative quantification(q RT-PCR)technique and physiological and biochemical indicators were used to determine the effects of exogenous NAA on the main protective enzymes activities,endogenous hormone content and related gene expression at three key stages of melon graft healing process.RNA-Seq technology was used to analyze the effects of exogenous NAA application on the main metabolic pathways and differentially expressed genes during the graft healing process,in order to systematically analyze the physiological and molecular mechanisms of exogenous NAA regulation of melon graft healing,in order to provide theoretical and practical basis for further improving grafting efficiency.The main findings are as follows:1.Effects of exogenous NAA application on the histological changes of the grafted melon grafted seedlings: The paraffin sections showed that the control plants formed isolated layer on the 2 DAG,and formed callus on the 6 DAG,and formed a new vascular bundle on the 9 DAG.Exogenous NAA treatment had no significant effect on the formation of the isolated layer during the graft healing process,but the formation period of callus and vascular bundle were 1 d earlier than the control.It indicated that exogenous NAA treatment can effectively promote the graft healing of oriental melon.2.Effects of exogenous NAA application on the activities of related enzymes and related genes during the healing process of oriental melon grafts: During the graft healing process,the activities of SOD and POD increased first and then decreased,reached the highest value at the stage of callus formation,and the enzymes activities were significantly higher than the control.The activities of PAL were higher than the control and reached the highest value at the formation of vascular bundle.The activities of PPO were significantly higher than the control at the formation of the isolated layer and indistinctively lower than the control with the grafting seedlings growth.Exogenous NAA treatment significantly increased the relative expression of Cm Fe-SOD,Cm Cu-Zn-SOD,Cm Mn-SOD and Cm PPO except Cm POD,and the trend of changes is basically consistent with the enzyme activities changes.It indicated that exogenous administration of NAA induced the expression of key enzymes genes that involved in graft union and increased the related enzymes activities,which promoted the graft healing process of grafting melon seedlings.3.Effects of exogenous NAA application on endogenous hormone content and auxin synthesis related genes: After exogenous application of NAA,except for GA3,the contents of IAA,ZR and ABA were significantly higher than the control at the formation of the callus and vascular bundles.The expression of Cm YUC10 was induced at the formation of the isolated layer,and the expression of Cm YUC4 and Cm YUC6 genes was induced at the formation of callus.The expression of Cm YUC8 in the whole process of graft healing was higher than the control,and it was significantly higher than the control at the formation of callus.It indicated that exogenous NAA significantly increased the content of phytohormone,and induced the expression of auxin synthesis gene during grafting healing,which promoting the development of callus and vascular bundle.4.Effects of exogenous NAA application on the transcriptome characteristics of oriental melon: 20996 genes were selected in the pumpkin and melon genomes.After selecting for differential genes,532 differential genes were selected in the control,and 4212 differential genes were selected by exogenous NAA.The GO and KEGG analysis of the differential genes revealed that the plant hormone signal transduction pathway and the lignin synthesis pathway were the most important in the melon and pumpkin genomes,and these two pathways were closely related to the graft healing process.After exogenous application of NAA,the expression of four hormones(IAA,ABA,GA3,ZR)signal transduction-related genes was higher than the control at the lately stage of grafting.The trend of change was consistent with the trend of hormone content.At the early stage of grafting,the expression level of the gene involved in the lignin synthesis is higher than the NAA treatment,but it is reversed at the lately stage of grafting.It indicated that the number of differential genes was increased after exogenous NAA application,and more metabolic activities were involved,which providing more material basis for the graft healing process.