Evaluation Of Core Germplasm And Study On Biosynthesis And Regulatory Mechanism Of Artemisinin In Artemisia Annua L.by Multiple Omics

Artemisinin is derived from Artemisia annua L.,a plant of the genus Artemisia,which is the first choice for the treatment of malaria.Artemisinin solves the problem of resistance of quinoline to Plasmodium falciparum and has become domestic and foreign.Recognized antimalarial drugs.In recent years,the global artemisinin dosage is about 180 tons,and it is mainly extracted from the leaves of A.annua.The artemisinin content in A.annua directly affects its extraction cost,which is the key to restrict the development of artemisinin downstream industry.Strengthening the collection and arrangement of resources,strengthening the cultivation of new varieties and standardized planting and promotion,in-depth study of the synthesis mechanism of artemisinin,and the efficient preparation of high-quality raw materials of artemisinin are the primary problems that need to be solved urgently.In this paper,the core germplasm resources of A.annua in China were collected and screened,and the genomic variation of 15 core germplasm was detected by whole genome resequencing,combined with full-length transcriptome sequencing,second-generation sequencing and LC-MS analysis to study the effects of light and plant hormones on artemisinin synthesis.The endogenous metabolites of two core germplasm with high-and low-artemisinin contents by broad-spectrum targeted metabolism technique,and the relationship between lncRNA and their differential characteristics were analyzed.The main results are as follows:1?Analysis of variation information of core germplasm collection of A.annua by re-sequencing technique.We collected wild and cultivated resources of A.annua from 27 provinces then established a germplasm bank,and selected 15 core producing areas for re-sequencing through resource evaluation.Fifteen samples produced 43.64-81.99G clean data,the comparison rate of all samples was between 98.49%and 100.00%,and the effective sequencing depth fluctuated between 22.44X and 38.67X.The whole genome variation site informations of A.annua from 15 different producing areas were obtained by re-sequencing technique,and the total amount of data reached 995G,and the database of gene variation polymorphism of Artemisia annua L.was constructed.2?Transcriptome sequencing revealed the molecular mechanism of light and hormone coordinately promoted the accumulation of Artemisinin in A.annua.Light and plant hormones are important external environmental factors,which can regulate some physiological processes of plants.In this paper,transcriptome and LC-MS were used to evaluate the effect of UV(Ultraviolet B Irradiation,UV-B)and gibberellin(Gibberellin,GA)on artemisinin synthesis.The results showed that the crosstalk between UV-B and GA induced artemisinin accumulation.A total of 14762 differentially expressed genes(DEGs)among different treatments were identified.UV-B and GA up-regulate the expression of ADS gene and CYP71AV1 gene,which are key genes of artemisinin biosynthesis,and promote the accumulation of artemisinin.According to the high expression level,84 coexpressed transcription factors were identified.Among them,MYB and NAC TFs are mainly involved in the regulation of artemisinin biosynthesis.Weighted gene coexpression network analysis showed that GA+UV in the blue module was positively correlated with artemisinin synthesis,and four candidate hub genes were associated with promoting artemisinin biosynthesis,in response to GA+UV treatment.3?Full-length transcriptome sequencing revealed the expression analysis of light treatment on alternative splicing of A.annua.Alternative splicing(Alternative splicing,AS)is extensive and complex in plants.We combinied the advantages of the long reading length of the third generation sequencing technology and the short reading length of the second generation sequencing technology,the full-length transcriptome database of A.annua was constructed,and a new set of alternative splicing data set of A.annua was obtained.A total of 11832 AS events were identified,of which 5854 were intron retention(Intron retention,IR).IR accounts for 48.48%of the total AS,indicating that the most common AS event is IR in A.annua.After blue light,red light and far-red light treatment,the number of up-regulated IR isoforms were more than the total number of down-regulated IR isoforms.Compared with white light and dark light,the number of IR isoforms up-regulated by blue light,red light and far red light was more than twice that of down-regulated IR isoforms.The results of GO enrichment showed that some genes had special functions and could produce many kinds of isoform.In the artemisinin biosynthesis pathway,there are 10 genes that produce 26 kinds of iso form,including 16 kinds of IR isoform,2 kinds of AA isoform and 8 kinds of other isoform.Therefore,the expression of IR isoform is regulated by blue light,red light and far-red light,and participates in the biosynthesis of artemisinin.4?Identification and prediction of lncRNA related to regulation of artemisinin biosynthesis.To explore the differences of broad-spectrum targeted metabolites of two core collections of YQ7(high-artemisinin)and JL1(low-artemisinin),combined with the transcriptome data,the relationship between lncRNA and the differences of the two chemical types was revealed.A total of 454 differential metabolites were identified by metabonomics,and 416 and 405 differential compounds were identified in YQ7 and JL1,respectively.There were significant differences in metabolic components between YQ7 and JL1.A total of 16 sesquiterpenes and 2 triterpenoids were identified from the existing terpenoid database.A total of 1419 novel lncRNA,were identified by transcriptome analysis.Among them,264 candidate lncRNA were identified,which laid a foundation for further exploration of the possible role of lncRNA in the regulation of artemisinin biosynthesis.In this study,the genetic information database of core germplasm of Artemisia annua was established,and the exogenous and endogenous factors of artemisinin synthesis were studied by multiple omics,which laid a foundation for revealing the mechanism of artemisinin synthesis and cultivating high-yielding lines of A.annua.