| Panax japonicus C.A.Meyer is a perennial herb of genus Panax in Araliaceae family.Its underground rhizome is the main medicinal part,which has both Panax ginseng nourishing and Panax notoginseng promoting blood circulation effects,hence it is known as the King of Herbs.For a long time,Chinese folk often dry it in the sun and then cook it into soup,which can not only be used as a food material,but also play a role in physical fitness.However,it is mainly used in pharmacology.It is often used as a medicinal material for the treatment of falls and injuries,as well as the auxiliary health care after the initial recovery of major diseases.The medicinal components of P.japonicus are mainly oleanane type pentacyclic triterpenoid saponins,which are white crystals,and have the effects of analgesia,tumor inhibition,immunity enhancement,anti-aging,refreshing and memory enhancement.Due to the high medicinal value of wild P.japonicus,it is endangered by people’s extensive excavation.It has very strict requirements on the growth environment and long growth cycle.It is a typical fat-loving shade plant with high humidity requirements,narrow ecological range and weak diffusion ability.Extreme climate leads to the extinction of this species at any time.It has now become one of the rare and endangered ‘seven Chinese herbal medicines’ in the country.In this study,the cultivated P.japonicus was treated with simulated natural drought,150 mmol/L and300 mmol/L salt stress,and the total saponins in the leaves,stems and rhizomes of P.japonicus were extracted and determined.Total RNA was extracted for transcriptome sequencing,and its transcriptome in response to drought and salt stress was analyzed to explore the correlation between total saponin content and gene expression after stress,and try to find key genes that can regulate saponin content after stress.The main results are as follows:1.Transcriptome sequencing of the leaves,stems and rhizomes of P.japonicus under drought and salt stress was carried out.After filtration,a total of 233.02 Gb clean data was obtained.After Trinity assembly,a total of 221804 Unigenes were obtained.Compare Unigenes with databases such as Nr,Uniprot,GO,COG and KEGG.There were 66488,35943,35475,13257 and 10839,respectively.Unigenes in KEGG were mapped to 91 pathways.Conducting differential expression gene screening analysis,it was found that the control group and drought treatment group obtained 102,63 and 124 differentially expressed genes in leaves,stems,and rhizomes,with 44,23 and 33 upregulated genes,58,36 and 86 downregulated genes;The results of the control group and 150 mmol/L Na Cl stress treatment showed that there were 769,568 and 480 differentially expressed genes in leaves,stems and rhizomes,respectively.Among them,245,161 and 149 genes were upregulated,524,407 and331 genes were downregulated;Under 300 mmol/L Na Cl stress,557,616 and 667 differentially expressed genes were obtained from leaves,stems and rhizomes,respectively.Among them,207,175 and 165 genes were upregulated,350,429 and 502 genes were downregulated.Under drought stress,the number of DEGs in P.japonicus showed rhizome > leaf > stem.Under low concentration salt stress,the number of DEGs obtained was leaf > stem > rhizome,while under high concentration salt stress,the number of DEGs was leaf < stem < rhizome.2.Under drought and salt stress,the total saponin content of P.japonicus showed an overall trend of rhizomes > leaves > stems.Under drought and 150 mmol/L Na Cl stress,the saponin content in the leaves of P.japonicus(2.87%,2.23%)and the total saponin content in the rhizomes(8.83%,5.67%)were increased compared to the control group’s leaves and rhizomes(1.97%,5.67%),indicating that drought and low concentration salt stress can accumulate saponins in P.japonicus,but the increase in drought stress is more significant.On the contrary,the total saponin content of P.japonicus under high concentration salt stress was lower than that of the control group.3.In this transcriptome sequencing,two metabolic pathways related to triterpenoid saponin synthesis were screened,namely,the terpenoid skeleton biosynthesis pathway(Ko00900)and the sesquiterpene and triterpene metabolism pathway(Ko009090).Compared with the drought group,the control group found that only three differentially expressed genes related to saponin synthesis were annotated in the rhizome,namely,Unigene187613,Unigene101529 and Unigene18274.Comparing the control group with the low concentration salt treatment group,it was found that differentially expressed genes related to saponin synthesis were released in the leaves,stems,and rhizomes,with Unigene40257 and Unigene205562 in the leaves;The stems include Unigene205562 and Unigene20949;The rhizomes include Unigene187613,Unigene205562,and Unigene42591.Comparing the control group with the high concentration salt treatment group,seven differentially expressed genes related to saponin synthesis were annotated in the rhizome,namely Unigene205562,Unigene187613,Unigene191116,Unigene22448,Unigene42591,Unigene101529,and Unigene57857.Finally,a correlation analysis was conducted between saponin content and the expression of these genes to obtain a key gene Unigene187613 that can induce the accumulation of saponin content in rhizomes under drought and salt stress conditions(β-Perfume resin synthase gene family,β-AS),two gene Unigene42591(isoprene synthase gene family,isps)and Unigene40257 were obtained,which have a positive regulation effect on the accumulation of saponin content under low concentration salt stress;Two genes that negatively regulate saponins under stress were obtained,namely Unigene101529(1-deoxy-D-ethylfructose-5-phosphate synthase gene family,DXS).It is worth noting that Unigene205562(SE)is expressed under salt stress conditions and also plays an important regulatory role in the response of P.japonicus to salt stress. |
