| Rosa roxburghii Tratt.has great development value due to its rich nutrition and effective substances.Its processed products cover food,medicine,health products,cosmetics,etc.,and which has become a key characteristic industry in Guizhou Province.Currently,R.roxburghii cultivated in production is mainly composed of several early approved varieties and recently selected genotypes,which are generally roughly distinguished by appearance traits.With the increasing demand for high-end,and specialized processing products of R.roxburghii,it is urgent to understand the nutritional and functional characteristics of different varieties or genotypes of fruit;at the same time,developing specific molecular markers for variety identification is also a necessary means of variety protection.This study was conducted using three varieties: ’Guinong 1’(Rr-1),’Guinong 5’(Rr-5),and ’Guinong 7’(Rr-7),two excellent genotypes: Guinong 3(Rr-3),Guinong 4(Rr-4),and a variant: R.roxburghii f.eseiosa Ku(Rrf).Comparative metabolomics was used to analyze the main metabolite composition of fruits,combined with antioxidant capacity and screening of chemical components of traditional Chinese medicine to explore characteristic metabolites;in the meantime,the In Del sites at the respective DNA level were screened through re-sequencing and specific molecular markers were developed for variety identification.The main findings are as follows:1.Quality performance and metabolome characteristics of R.roxburghii fruits The basic quality indicators of the fruits of the six genotypes showed differences,with Rr-1 having the largest fruit type index,Rr-5 having the largest single fruit weight and the highest vitamin C content,Rr-7 containing the highest total phenols,total flavonoids,and total triterpenes showing the highest antioxidant capacity,and Rr-4 having the highest lipid content.By using ultra high performance liquid chromatography/tandem mass spectrometry(UPLC-ESIMS/MS)to identify and quantify the composition of metabolites in six R.roxburghii genotypes,a total of 723 secondary metabolites were identified,mainly including phenolic acids,terpenes,flavonoids,amino acids and their derivatives,nucleotides and their derivatives,tannins,lipids,lignans and coumarins,alkaloids,organic acids,and others.Through the traditional Chinese medicine system pharmacology database and analysis platform(TCMSP),191 chemical components and 44 antioxidant substances were selected,mostly phenolic acids,flavonoids,and triterpenoid secondary metabolites.Comparative analysis of fruit metabolisms between R.roxburghii and some Rosaceae fruit trees showed that only 293 specific secondary metabolites were detected in R.roxburghii fruit,and most of them were phenolic substances.The above results provided a sufficient basis for explaining the medical and health value of R.roxburghii.2.Characteristic metabolites mining By comparing the differences in metabolites among genotypes,the characteristic metabolites of each genotype were selected based on the criteria that a component was unique or exhibited significant differences in content compared to other genotypes.There were 14 characteristic metabolites in Rr-1,and the phenolic acid metabolites were the most,with only the relative quantitative up-regulation of 3,5-ODicaffeoylquinic Acid Methyl Ester and 3,4-Dihydroxy-L-phenylalanine;Rr-3 had only three characteristic metabolites,3,4-Dihydroxy-L-phenylalanine,Quercetin-5-O-glucuronide and2,6-Dimethoxyhydroquinone-1-O-glucoside,and only Quercetin-5-O-glucuronide has a relatively quantitative up-regulation;There were 49 characteristic metabolites in Rr-4,most of which were lipids,and multiple lysophosphatidylcholine isomers were relatively quantitatively up-regulated;Rr-5 had only four characteristic metabolites,namely Methyleugenol,Roxburic Acid,Quercetin-5-O-glucuronide and Muconic acid;Rr-7 had 55 characteristic metabolites,with the largest number of secondary metabolites of amino acids,followed by phenolic acids,and among them,the relative quantitation of p-Coumarylmalic acid,4-Methoxycinnamic acid,1-(2,4,5-Trimethoxyphenyl)-1,2-propanedione,3,4-Digalloylshikimic acid,Syringalide A,6-O-Feruloyl-D-glucose,Sinapoyl-pcoumaroyltartaric cid,1-O-p-Coumaroylquinic acid,Feruloylferuloyltartaric acid,4-Hydroxybenzaldehyde,p-Coumaric acid-4-O-glucoside showed an upward trend compared with other genotypes;Rr-f has 23 characteristic metabolites,mainly including phenolic acid,organic acid and triterpenoid secondary metabolites,and the relative quantitation of the only three terpene metabolites,Roxburic Acid,1β,2α,3α,19α,23-Pentahydroxyurs12-en-28-oic acid,27,28-Dicarboxyl ursolic acid were up-regulated in Rr-f.The results pointed out the direction for the specific development and utilization of different varieties(genotypes).3.Development of specific molecular markers A total of 668029 In Del loci were screened using the re-sequencing data from three genotypes of R.roxburghii.Some specific In Del loci of each genotype were randomly selected to establish 5 genotype specific markers.Total 174 pairs of primers were designed.Four sets of specific primer pairs were selected to successfully identify 3 varieties,Rr-1,Rr-5,Rr-7,Rr-f.Therefore,a molecular identification method for the main varieties of R.roxburghii was established. |
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