Effects Of Exogenous Methyl Jasmonate On Transcriptome And Micro Rnaome Of Euphorbia Kansui

Euphorbia kansui Liou is one of the unique traditional Chinese medicine.The pharmacological study showed that the main component of E.kansui was terpenoids,which has activities of anti-tumor,anti-cancer,anti-virus,anti-oxidant,anti-cell division and regulating immune system.The jasmonate is an important growth regulator in plants,which can mediate diverse physiological processes,such as plant growth and development,as well as the accumulation of secondary metabolites.Some studies have reported that the effect of MeJA on biosynthesis of the secondary metabolites is more significant than salicylic acid(SA)and nitric oxide(NO).Illumina sequencing technology can be used to obtain the information of all mRNAs and miRNAs in living organisms,comprehensive data of analysis about differentially expressed genes and miRNAs,the mining of new genes and novel miRNAs,gene function annotation,the prediction of miRNAs target mRNAs and the annotation of function.All these data can help us to further understand many biological problems.This paper used sequencing techniques of transcriptome and micro RNAome to study the effect of exogenous 0.5 mg/L MeJA on development and regulatory of the secondary metabolites biosynthesis in E.kansui.Through the studies above,it was hoped to provide theoretical basis for further study of response mechanism of kansui to exogenous MeJA,and also provide basic data for genetic engineering research of green development of E.kansui.The main research results were as follows:1.By assembling high quality reads obtained from each transcriptome group,96 040 unigenes were obtained,among which 52 142,40 079,36 333,20761 and 38 008 unigenes were annotated in Nr,Swiss-Prot,COG,KEGG and GO databases,respectively.To identify the DEGs,the samples of four groups were pairwise compared by using the edgeR package.The DEGs were annotated with the GO function classification,and pathway significant enrichment was carried out using KEGG pathway as the unit,the significant enrichment pathways in DEGs were found through hypergeometric test.In addition,RNA expression patterns of the DEGs in four groups were directly compared by Short Timeseries Expression Miner(STEM)cluster,and 7 significant gene expression profiles were identified.2.The high-throughput sequencing was performed to obtain the miRNAs transcriptome dadabase.The target mRNAs of miRNAs were predicted by software patmatch.In total,875 conserved miRNAs corresponding to 11 277 target mRNAs were identified in 8 samples,among which 168 known miRNAs represented 6 019 target mRNAs.In known miRNAs,the most abundant expression was the MIR169 family,followed by the MIR156,MIR166,MIR171 and MIR395 family.This study also identified 707 novel miRNAs and their corresponding 5 258 target mRNAs.In addition,the expression patterns of DEMs were predicted by STEM cluster,and 4 were significant miRNAs expression profiles.According to the correlation analysis between miRNAs and mRNAs,452 miRNA-mRNA pairs were identified to present an anti-correlationship.3.Through analysis of DEGs in the terpenoid biosynthesis pathway,18 key unigenes were selected for qPCR verification,the verification results were basically consistent with the transcriptome results.Based on the analysis of miRNAs in the terpenoid biosynthesis pathway,33 miRNA-mRNA pairs were correlated with terpenoid biosynthesis.7 DEMs were selected for qPCR verification,the verification results were basically consistent with the results of miRNAs sequencing.Furthermore,taking euphadienol as standard substance,the content of the total terpenoids and euphadienol in four groups were detected by UV-visible spectrophotometer and high performance liquid chromatography(HPLC).The results showed that the content of the total terpenoids and euphadienol were increased after 24 hours of MeJA treatment,and the content of the total terpenoids and euphadienol were highest at 36 hours of MeJA treatment.In summary,exogenous MeJA maintains the homeostasis physiological and metabolic activities of plant by affecting the expression patterns of kansui unigenes.Mi RNAs participate in the response of plants to exogenous MeJA by negatively regulating their target mRNAs.Exogenous MeJA promoted the expression of unigenes in the MVA pathway of terpenoid biosynthesis,thereby enhanced the content of the total terpenoids and euphadienol.Mi RNAs participated in the regulation of terpenoid biosynthesis by negatively regulating their target mRNAs at the early stage of MeJA treatment.The MVA pathway and the MEP pathway of terpenoid skeleton biosynthesis may be negatively regulated each other in response to the treatment of exogenous MeJA,but this speculation still needs to be verified by further molecular biological research.