| China is the largest producer and importer of kiwifruit,while the kiwifruit canker caused by Pseudomonas syringae pv.actinidiae(Psa)has seriously affected the development of kiwifruit industry in China.At present,the prevention of kiwifruit bacterial canker disease is still the main treatment,and there is no particularly effective treatment.The wide variety of plant species hinders the development of disease resistance breeding of ornamental plants,but breeding resistant varieties is the most effective and lasting method to control pathogens.In order to clearly study the anti-canker mechanism of kiwifruit and provide molecular basis for kiwifruit canker breeding.In this study,kiwifruit "Xuxiang" sensitive to canker and "Donghong" resistant to canker infected by Pseudomonas syringae pv.actinidiae for 0 h,12 h and 24 h were used to construct a chain-specific library for sequencing,and then build a personalized analysis pipeline to explore the mechanism of kiwifruit canker-resistant disease.The main findings are as follows:(1)Establish a personalized pipeline to analyze the screening differential genes of kiwifruit under Psa stress: The close evolutionary relationship between Actinidia chinensis var.deliciosa and Actinidia chinensis var.chinensis was demonstrated by non-reference transcription analysis.The GO and KEGG enrichments of differential genes indicate the plant immunity,phytoalexin metabolism,flavonoid metabolism,multiple plant hormone metabolism,stomatal movement,cell wall metabolism and other processes or pathways related to plant defense against bioyic stress.3342 lnc RNAs were assembled and identified with a reliability of 2 to 4.According to the RT-q PCR results,it is speculated that the protein coding genes Acc15563,Acc31327 and Acc11580 and the conservative lnc RNA genes MSTRG.5146.1,MSTRG.30577.2,MSTRG.14166.1,MSTRG.10141.1 and MSTRG.26040.3 may be involved in the resistance of Psa stress in kiwifruit.(2)Establish the pipeline of pathogenic bacteria transcriptome in the interaction transcriptome: Analyze the differentially expressed genes in the Psa transcriptome by using the reads not mapping to Red5 genome,which provides the basis for finding Psa effector factors.(3)Establish WGCNA analysis pipeline of kiwifruit protein encoding gene under Psa stress: The expression matrix of all genes in kiwifruit are divided into 22 modules,and the blue module related to resistance to kiwifruit canker is identified.The genes in blue module are performanced GO and KEGG enrichment enrichments,and the processes or pathways related to biological stress are identified.It was proved that WGCNA was feasible for the analysis of the resistance mechanism of kiwifruit canker disease.According to the RT-q PCR results,top ten genes with most edges in the Blue module may be involved in the resistance of Psa stress in kiwifruit.(4)Exploration results of Ac HD-Zip gene family expression in transcriptome of kiwifruit under Psa biotic stress: 77 Ac HD-Zip genes are identified in Red5 genome,and the gene structure and protein sequence in each subfamily are conservative.And these 77 Ac HD-Zip genes expressed differently in the resistant material "Xuxiang" and the sensitive material "Donghong",which may play a role in the process of resistance to Psa stress of kiwifruit.According to the RT-q PCR results,it is speculated that Acc06400、Acc25123 and Acc22265 may be involved in the resistance of Psa stress in kiwifruit.(5)Developed a R package and some scripts: Developed personalized scripts()and an R package Bio Uncle()to simplify the established analysis pipeline and provide a basis for future research.Based on the transcriptome data of kiwifruit materials resistant and sensitive to Psa,this study obtained the biological processes and pathways resistant to kiwifruit canker disease,established the analysis pipelinse of Dual RNA-seq,WGCNA and HD-Zip gene family,and developed personalized scripts and R package Bio Uncle to simplify the process,providing a basis for future research. |
