Screening For Regulation Elicitor Of Secondary Metabolism Pathways And Transcriptome Studies Of Astragalus Membranaceus (Fisch.)Bge.

Astragalus membranaceus (Fisch.) Bge. is an important medicinal plant grown in North China, which are widely used in clinical formula, has cardiotonic effect, against myocardial ischemia and improve immunity, reduce blood glucose, anti-aging, protect viscera and so on, many kinds of pharmacological action. Flavones, polysaccharides and saponins are major medicinal effective ingredients in Astragalus membranaceus. These biological active ingredients belong to secondary metabolites. Now the secondary metabolite production directly from the extraction plants of Astragalus. However, one of the major obstacles is the low yield of plant secondary metabolites in Astragulus. Due to limited genomic and transcriptomic data, the biosynthetic pathway of secondary metabolite, the major bioactive compound in astragalus membranaceus, is currently unclear. Identification of candidate genes and crucial regulators of gene expression involved in the secondary metabolite biosynthetic pathway will significantly contribute to the understanding of the biosynthetic pathway of secondary metabolite.In order to make the secondary metabolic pathways of Astragalus from the background of normal metabolism, and was catch to study. This study using the orthogonal experiment design, polysaccharide, saponin and flavonoid content of Astragalus were measure, screening for regulation elicitor of secondary metabolism pathways. Orthogonal test results showed that the silver nitrate and potato culture medium solution were the best elicitor of polysaccharide metabolic pathways, the best processing part was root, the optimum processing time was6-9days, and best detection part was root. The methyl jasmonic acid solution and endophytic fluid was the best elicitor of saponins secondary metabolic pathways, the best processing part was root, the optimum processing time was3days, and best detection part was root. The silver nitrate solution was the best elicitor of flavones secondary metabolic pathways, the best processing part was root, the optimum processing time was9days, and best detection part is leaf.At the same time, used454/Roche GS FLX sequencing technology, root, stem and leaf of Astragalus were sequenced1/2plate run de novo mix of total RNA proportion.This1/2run produced742721high quality reads with an average length413bases. Of the high quality reads,86%contained were used for assembly, N50length was1205bases. We got9893unigene through blast cluster. All unigene sequences were compared in the three major public protein databases using the BLASTX algorithm, total of9732unigene sequences were annotated, which accounted for 98.37%of the total unigene sequences. Functional classification and pathway assignment was performed by the Kyoto Encyclopedia of Genes andGenomes (KEGG). Of these unigene sequences,2877had significant matches in the database sequences to article45of metabolic pathways. Among those,36unigene sequences were assigned to Glycan Biosynthesis and Metabolism of metabolic pathways,68unigene sequences were assigned to Metabolism of Terpenoids and Polyketides of metabolic pathways.A total of12880isotig sequences contained1729SSRs, average distance was7.97kb. Six types of mononucleotide to hexanucleotide,127kinds repeat motif occurrenced. The genome of Astragalus membranaceus had not been sequenced yet, nevertheless it is possible to generate gene model predictions using the Astragalus membranaceus transcriptome and the genomes of Glycine max, Medicago truncatula or Lotus japonicus, which are the three closest related species that have genome sequence available. Transcriptome was mapped to the Medicago truncatula genome, representing2039gene loci and21727transcript models, identity90.14%. Transcriptome was mapped to the Glycine max genome, representing1948gene loci and36864transcript models, identity90.40%. Transcriptome was mapped to the Lotus japonicus genome, representing1003gene loci and10056transcript models, identity89.77%. Alternative splicing site types are290to predict using the genomes of Medicago truncatula,284to predict using the genomes of Glycine max,273to predict using the genomes of Lotus japonicas. AG<exon>GT,<L-Gap> and <R-Gap> types were the most commonly found in the three leguminous model plants.This study on the screening of elicito secondary metabolism pathways and transcriptome studies of Astragalus membranaceus, obtained the elicitor of corresponding metabolic pathway and the key regulation genes and transcription factors. The study provided theoretical quidance and technical support for controling secondary metabolic pathway through genetic engineeting means and improving the the content of astragalus secondary metabolites qualitative.