The Preliminary Study Of CircRNA Which Is Closely Related To Tomato Late Blight

CircRNA is a class of endogenous noncoding RNAs that are ubiquitous in organisms.Inrecent years,studies have shown that circ RNA can adsorb mi RNA and indirectly regulate gene expression,and mi RNAs have been fully confirmed to act on m RNAs to play an important regulatory role in plant growth and resistance to biological and abiotic stresses.Tomato is a kind of dicotyledonous model plant which is widely planted in the world.The late blight caused by Phytophthora infestans brings great economic loss to tomato production.In addition,there is currently a lack of reports on the circ RNA associated with tomato disease,and the analysis of its functional properties,especially the study of circ RNA-mi RNA-m RNA regulatory networks,is not yet systematic.In this study,the circ RNA closely related to diseases in tomato was discovered,and the related circ RNA functions were preliminary explored through experiments.The specific research contents are as follows:Firstly,68 related circ RNA were discovered by analyzing the transcriptome database constructed in the early stage of the laboratory.There were 27 down-regulated circ RNA and 9up-regulated circ RNA.By combining the gene expression values with the GO functional annotation results of the parental genes,two exon circ RNA closely related to late blight of tomato were obtained.After that,a relationship network of Ecirc RNA-mi RNA was constructed.Based on the analysis of target genes of 11 mi RNA,two disease-related mi RNA were selected as follow-up targets.They were mi R477-3p and mi R9478-5p.Secondly,Ecirc RNA45 and Ecirc RNA47 were selected as the focus of this study.They were verified by PCR with forward and reverse primers,and their expression characteristics were analyzed by real-time quantitative PCR.It was found that their expression levels in leaves were higher than those in roots and stems,and they all responded to Phytophthora infestans and were negatively correlated with the expression of mi R477-3p and mi R9478-5p.In addition,it was found that the expression of mi R477-3p and mi R9478-5p were down-regulated,while their target genes were up-regulated,indicating that circ RNA could indirectly regulate the expression of genes by adsorbing mi RNA in tomato response to stress.Finally,In order to further explore the regulatory relationship of circ RNA-mi RNA,I cloned Ecirc RNA45,Ecirc RNA47,MIR477-3p and MIR9478-5p in tomato and successfully constructed the corresponding overexpression vector and established four transient expression systems.In tomato leaves of over-expressing Ecirc RNA45 and Ecirc RNA47,the expression levels of mi R477-3p and mi R9478-5p were significantly decreased and the target genes were significantly up-regulated.In tomato leaves of over-expressing MIR477-3p and MIR9478-5p,the target genes were down-regulated.It was shown that Ecirc RNA45 and Ecirc RNA47 can indirectly regulate m RNA expression by silencing mi RNA,and thus play a role in plants and pathogens.In summary,this study used the combination of bioinformatics analysis and experiments to predict,mine,and identify circ RNA that were closely related to disease in tomato,and performed related functional analysis,which deepened the interaction between plants and pathogens.The understanding of the circ RNA mechanism.