The Nutritional Omics Study Of Akebia Trifoliata And Isolation And Identification Of Pathogen Bacteria From It

As a climbing perennial deciduous vine of the Akebia family,Akebia trifoliata(Thunb.)Koidz.(Hereafter referred as A.trifoliata)has been used as medicine and wild fruit for more than one thousand years in China.The lack of chloroplast genome,full-length transcripts and metabolomics in fruit of A.trifoliata fruit seriously hinders the research of A.trifoliata in the field of food and molecular biology.Moreover,as a new economic crop,A.trifoliata is severely threatened by fruit diseases caused by microorganisms with the expansion of planting area.Therefore,it is of great significance to study the related omics and pathogen bacteria of A.trifoliata.The chloroplast genome of A.trifoliata was sequenced and assembled for the first time,and the evolutionary relationship was analyzed.The results showed that the gene was 157,952 bp in size,encoding 131 genes,containing large single copy region of 86,596 bp,small single copy region of 19,058 bp,and reverse repeat sequence of26,149 bp,with GC content of 38.70%.The phylogenetic results showed that A.trifoliata was closely related to Akebia quinata,which was consistent with the taxonomic status of all kinds of Akebia.At the same time,the roots,stems,leaves,flowers and fruits of A.trifoliata at different development stages were taken as samples to sequence and annotate the full-length transcripts of A.trifoliata.A total of1,414,525 transcripts were obtained,with a total library size of 338.43 GB.GC content was 40.42%.There were 285,947 transcripts that could be annotated in the common function database,accounting for 20.22%.The prediction and family classification of transcripts encoding transcription factors will provide references for further research on fruit development regulating genes and accumulation of fruit nutritive metabolites.In order to study nutrient metabolites of A.trifoliata fruits,the peels,pulp and seeds were taken as samples to make non-targeted metabolomics analysis.A total of10,772 and 7,149 positive and negative ions were obtained.By analyzing the differential metabolites in different tissues,the metabolic differences and functional compounds were further identified.According to the quantitative analysis of metabolites,among the top 100 metabolites,the pulp and seeds were rich in phenylalanine,isoleucine(top 10)and other essential amino acids.The content of uronic acid in pulp was relatively high,which was also one of the contributing components of pulp sweetness.The seed is rich in essential unsaturated fatty acids such as oleic acid,monolenic acid,palmitic acid,citric acid and glycerol.In addition,the analysis also showed that the contents of functional metabolites were rich,such as choline,ursolic acid,chlorogenic acid,soya saponin and ganoderic acid.According to the metabonomics,the nutritive value is high,so the study on its pathogen bacteria is of practical significance.Through isolating and identifying the pathogen bacteria that made the fruit shrink and blacken,four fungi,pestalotiopsis oryzae,colletotrichum nymphaeae,Nigrospora sphaerica and cladosporium asperulatum were obtained.The isolated fungus N.sphaerica which can cause the fruit shrink and blacken to death when grafted to holboellia latifolia wall was identified as N.sphaerica by morphology and molecular biology.At the same time,the biological control of pathogen bacteria was preliminarily studied.The Bacillus subtilis had strong bacteriostatic effect.The impact of N.sphaerica causing the fruit shrink and black to death was first reported.The genome of N.sphaerica was sequenced,assembled and annotated.The genome size of N.sphaerica was 51.75 Mb,which was close to the results of genome survey(50.55Mb).The heterozygosity rate was about 0.22%,and the number of repeats was about 15.96%.The results of functional annotation showed that the number of genes with functional annotation was 10,106,accounting for 87.74%.The interaction between N.sphaerica and A.trifoliata fruits was analyzed by transcriptome analysis.A total of 11,253 genes were detected and 202 new genes were predicted.Intertranscriptome analysis of the genome of N.sphaerica and its alignment gene set showed an average alignment rate of 24.43% and 22.19%.Compared with 12-hour samples,718 different genes were obtained,which were involved in fungal pathogenicity and fruit defense response.Through the isolation and identification of pathogen bacteria of A.trifoliata,it provides a reference for the prevention and treatment of diseases caused by A.trifoliata fungus.A large number of gene structure and function annotation data obtained from the transcriptome of pathogen-host interactions of bacteria have laid a certain foundation for the further researches on key genes related to disease and disease resistance,prevention and treatment.