This study investigates the genetic diversity and evolutionary relationships of different Artemisia argyi germplasm resources to provide a basis for germplasm identification, variety selection, and resource protection. A total of 192 germplasm resources of A. argyi were studied, and EST-based simple sequence repeat(EST-SSR) primers were designed based on transcriptomic data of A. argyi. Polymerase chain reaction(PCR) amplification was performed on these resources, followed by fluorescence capillary electrophoresis to detect genetic diversity and construct DNA fingerprints. From 197 pairs of primers designed, 28 pairs with polymorphic and clear bands were selected. A total of 278 alleles were detected, with an average of 9.900 0 alleles per primer pair and an average effective number of alleles of 1.407 2. The Shannon's diversity index(I) for the A. argyi germplasm resources ranged from 0.148 1 to 0.418 0, with an average of 0.255 7. The polymorphism information content(PIC) ranged from 0.454 5 to 0.878 0, with an average of 0.766 9, showing high polymorphism. Cluster analysis divided the A. argyi germplasm resources into three major groups: Group Ⅰ contained 136 germplasm samples, Group Ⅱ contained 45, and Group Ⅲ contained 11. Principal component analysis also divided the resources into three groups, which was generally consistent with the clustering results. Mantel test results showed that the genetic variation in A. argyi populations was to some extent influenced by geographic distance, but the effect was minimal. Structure analysis showed that 190 germplasm materials had Q≥ 0.6, indicating that these germplasm materials had a relatively homogeneous genetic origin. Furthermore, 8 core primer pairs were selected from the 28 designed primers, which could distinguish various germplasm types. Using these 8 core primers, DNA fingerprints for the 192 A. argyi germplasm resources were successfully constructed. EST-SSR molecular markers can be used to study the genetic diversity and phylogenetic relationships of A. argyi, providing theoretical support for the identification and molecular-assisted breeding of A. argyi germplasm resources.
Artemisia/classification* ; Microsatellite Repeats ; Genetic Variation ; Expressed Sequence Tags ; DNA Fingerprinting ; Phylogeny ; Polymorphism, Genetic ; DNA, Plant/genetics* ; Genetic Markers

Two novel skeleton sesquiterpenoids (1 and 6), along with four new iphionane-type sesquiterpenes (2-5) and six new cyperane-type sesquiterpenes (7-11), were isolated from the whole plant of Artemisia hedinii (A. hedinii). The two novel skeleton compounds (1 and 6) were derived from the decarbonization of iphionane and cyperane-type sesquiterpenes, respectively. Their structures were elucidated through a comprehensive analysis of spectroscopic data, including high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) and 1D and 2D nuclear magnetic resonance (NMR) spectra. The absolute configurations were determined using electronic circular dichroism (ECD) spectra, single-crystal X-ray crystallographic analyses, time-dependent density functional theory (TDDFT) ECD calculation, density functional theory (DFT) NMR calculations, and biomimetic syntheses. The biomimetic syntheses of the two novel skeletons (1 and 6) were inspired by potential biogenetic pathways, utilizing a predominant eudesmane-type sesquiterpene (A) in A. hedinii as the substrate. All compounds were evaluated in LX-2 cells for their anti-hepatic fibrosis activity. Compounds 2, 8, and 10 exhibited significant activity in downregulating the expression of α-smooth muscle actin (α-SMA), a protein involved in hepatic fibrosis.
Artemisia/chemistry* ; Sesquiterpenes/chemical synthesis* ; Molecular Structure ; Humans ; Liver Cirrhosis/genetics* ; Biomimetics ; Plant Extracts/pharmacology*

Artemisiae Scoporiae Herba is derived from Artemisia scoparia or A. capillaris. The accurate identification of the herbs, particularly when dealing with bulk samples, is critical for ensuring the quality and efficacy of the medicinal product. This study aimed to establish a comprehensive molecular approach by combining multiple markers for the precise identification of Artemisiae Scoporiae Herba. The ITS2 from A. scoparia, A. capillaris, and other common Artemisia species were retrieved from GenBank. MEGA was used to build a phylogenetic tree with these sequences, and the effectiveness of ITS2 in species identification was assessed. The analysis revealed that while ITS2 could distinguish Artemisiae Scoporiae Herba from other closely related species of Artemisia, it was insufficient to differentiate between A. scoparia and A. capillaris. To address this limitation, the chloroplast genome of A. capillaris was assembled and compared with the published chloroplast genomes of A. scoparia and A. capillaris, on the basis of which a DNA mini-barcode was developed. The rpoA-rps11 region was selected as the target for the development of mini-barcode due to its potential for distinguishing between these two species. Specific primers were designed to differentiate A. scoparia from A. capillaris. The ITS2 sequences and the newly developed mini-barcode were used together for Sanger sequencing to identify individual samples of Artemisiae Scoporiae Herba, while DNA metabarcoding was employed for the identification of bulk samples. The identification results of representative individual samples and bulk samples from different regions consistently confirmed A. capillaris. This study established a method that combined ITS2 and mini-barcode to identify bulk samples of Artemisiae Scoporiae Herba from different regions. This approach overcomes the limitations of morphological and chemical methods, enhancing species identification accuracy and supporting a stable supply of medicinal materials.
Artemisia/classification* ; DNA Barcoding, Taxonomic/methods* ; Phylogeny ; DNA, Plant/genetics* ; DNA, Ribosomal Spacer/genetics*

Artemisia argyi is an important medicinal and economic plant in China, with the effects of warming channels, dispersing cold, and relieving pain, inflammation, and allergy. The essential oil of this plant is rich in volatile terpenoids and widely used in moxi-bustion and healthcare products, with huge market potential. The bZIP transcription factors compose a large family in plants and are involved in the regulation of plant growth and development, stress response, and biosynthesis of secondary metabolites such as terpenoids. However, little is known about the bZIPs and their roles in A. argyi. In this study, the bZIP transcription factors in the genome of A. argyi were systematically identified, and their physicochemical properties, phylogenetic relationship, conserved motifs, and promoter-binding elements were analyzed. Candidate AarbZIP genes involved in terpenoid biosynthesis were screened out. The results showed that a total of 156 AarbZIP transcription factors were identified at the genomic level, with the lengths of 99-618 aa, the molecular weights of 11.7-67.8 kDa, and the theoretical isoelectric points of 4.56-10.16. According to the classification of bZIPs in Arabidopsis thaliana, the 156 AarbZIPs were classified into 12 subfamilies, and the members in the same subfamily had similar conserved motifs. The cis-acting elements of promoters showed that AarbZIP genes were possibly involved in light and hormonal pathways. Five AarbZIP genes that may be involved in the regulation of terpenoid biosynthesis were screened out by homologous alignment and phylogenetic analysis. The qRT-PCR results showed that the expression levels of the five AarbZIP genes varied significantly in different tissues of A. argyi. Specifically, AarbZIP29 and AarbZIP55 were highly expressed in the leaves and AarbZIP81, AarbZIP130, and AarbZIP150 in the flower buds. This study lays a foundation for the functional study of bZIP genes and their regulatory roles in the terpenoid biosynthesis in A. argyi.
Gene Expression Profiling ; Phylogeny ; Artemisia/genetics* ; Basic-Leucine Zipper Transcription Factors/metabolism* ; Terpenes ; Gene Expression Regulation, Plant

Artemisia stolonifera is a relative of A. argyi. The two species are difficult to be distinguished due to the similarity in leaf shape and have even less distinctive features after processing. This study aims to establish a method to quickly distinguish between them. At the same time, we examined the reasonability and applicability of the specific polymerase chain reaction(PCR) method. The C/T single nucleotide polymorphism was detected at the position 202 of the sequence, based on which specific primers were designed to identify these two species. The PCR with the specific primer JNC-F and the universal primer ITS3R produced a specific band at 218 bp for A. argyi and no band for A. stolonifera, which can be used to detect at least 3% of A. argyi samples mixed in A. stolonifera samples. The PCR with the specific primer KY-F and the universal primer ITS3R produced a specific band at 218 bp for A. stolonifera and no band for A. argyi, which can be used to detect at least 5% of A. stolonifera samples mixed with A. argyi. The limit of detection of the established method was 5 ng DNA. The established PCR method can accurately distinguish between A. stolonifera and A. argyi, which provides an experimental basis for the quality control of A. stolonifera and determines whether the herbs are adulterated.
Artemisia/genetics* ; Trichomes ; Polymerase Chain Reaction ; Nucleic Acid Amplification Techniques ; Plant Leaves/genetics*

Artemisia Argyi Folium, a traditional Chinese medicine of important medicinal and economic value, sees increasing demand in medicinal and moxibustion product market. Screening stable and reliable reference genes for quantitative real-time PCR(qRT-PCR) is a prerequisite for the analysis of gene expression in Artemisia argyi. In this study, eight commonly used reference genes, Actin, 18s, EF-1α, GAPDH, SAND, PAL, TUA, and TUB, from the transcriptome of A. argyi, were selected as candidate genes. The expression of each gene in different tissues(roots, stems, and leaves) of A. argyi and in leaves of A. argyi after treatment with methyl jasmonate(MeJA) for different time(0, 4, 8, 12 h) was detected by qRT-PCR. Then, geNorm, NormFinder, BestKeeper, ΔCT, and RefFinder were employed to evaluate their expression stability. The results demonstrated that Actin was the most stable reference gene in different tissues and in leaves treated with MeJA, and coming in the second was SAND. Furthermore, the expression of DXS and MCT which are involved in terpenoid backbone biosynthesis was detected in different tissues and after MeJA treatment. The results showed that the expression patterns of DXS and MCT in different tissues and under MeJA treatment calculated with Actin and SAND as internal reference genes were consistent, which validated the screening results. In conclusion, Actin is the most suitable reference gene for the analysis of gene expression in different tissues of A. argyi and after MeJA treatment. This study provides valuable information for gene expression analysis in A. argyi and lays a foundation for further research on molecular mechanism of quality formation of Artemisia Argyi Folium.
Artemisia/genetics* ; Gene Expression Profiling ; Genes, Plant/genetics* ; Plant Leaves/genetics* ; Real-Time Polymerase Chain Reaction ; Reference Standards ; Transcriptome

To clarify the content characteristics of mineral elements in different Artemisia argyi germplasm resources and their relationship with the quality properties of Artemisiae Argyi Folium, this study measured the content of 10 mineral elements including nitrogen(N), phosphorus(P), potassium(K), calcium(Ca), magnesium(Mg), aluminum(Al), manganese(Mn), iron(Fe), copper(Cu), and zinc(Zn) in 100 Artemisia argyi germplasm samples. Besides, their relationship with the quality properties of Artemisiae Argyi Folium was explored by correlation analysis, path analysis, and cluster analysis. The results demonstrated that the variation coefficient of the 10 mineral elements in Artemisiae Argyi Folium ranged from 12.23% to 64.38%, and the genetic diversity index from 0.97 to 3.09. The genetic diversities of N, P, and Zn were obvious. As revealed by the correlation analysis, N, P, and K showed strong positive correlations with each other. Except that Mg and Al were negatively correlated, Ca, Mg, Al, Mn, Fe, Cu, and Zn were positively correlated. The correlation analysis of mineral elements with the quality properties of Artemisiae Argyi Folium proved the significant correlations of 17 pairs of characters. According to the path analysis, P, K, Ca, and Mn greatly affected the yield of Artemisiae Argyi Folium, P, K, and Mg the output rate of moxa, N, P, and K the content of total volatile oil, P and K the content of eucalyptol, and P, K, and Ca the content of eupatilin. The 100 germplasm samples were clustered into three groups. Specifically, in cluster Ⅰ, the enrichment capacity of P, K, and Mg elements was strong, and the comprehensive properties of mineral elements were better, implying good development potential. Ca, Mn, Fe, and Zn elements in cluster Ⅱ and N and Al in cluster Ⅲ displayed strong enrichment capacities. This study has provided new ideas for resource evaluation and variety breeding of A. argyi and also reference for fertilizer application.
Artemisia/genetics* ; Iron ; Minerals/analysis* ; Plant Breeding ; Plant Leaves/chemistry*

Artemisia indica is an important medicinal plant in the Asteraceae family, but its molecular genetic information has been rarely reported. In this study, the chloroplast genome of A. indica was sequenced, assembled, and annotated by the high-throughput sequencing technology, and its sequence characteristics, repeat sequences, codon usage bias, and phylogeny were analyzed. The results showed that the length of the chloroplast genome for A. indica was 151 161 bp, which was a typical circular four-segment structure, including two inverted repeat regions(IRs), a large single-copy(LSC) region, and a small single-copy(SSC) region, with a GC content of 37.47%. A total of 132 genes were annotated, and 114 were obtained after de-duplication, including 80 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. Fifty long repeat sequences and 191 SSRs were detected in the chloroplast genome of A. indica, and SSRs were mainly single nucleotides. Codon usage bias analysis showed that leucine was the most frequently used amino acid(10.77%) in the chloroplast genome, and there were 30 codons with relative synonymous codon usage(RSCU)>1 and all ended with A/U. The phylogenetic tree constructed based on the chloroplast genomes of the 19 species from the Asteraceae family showed that A. indica and A. argyi were closest in the genetic relationship, and Artemisia species clustered into separate evolutionary branches. The results of this study are expected to provide a theoretical basis for the genetic diversity and resource conservation of Artemisia medicinal plants.
Genome, Chloroplast ; Phylogeny ; Artemisia/genetics* ; Codon/genetics* ; Base Composition ; Plants, Medicinal/genetics*

In this study, in order to evaluate the phenotypic diversity of Artemisia argyi germplasm resources and improve its efficiency of cultivation and breeding, 100 accessions of A. argyi germplasm resources from 58 regions in China were collected, 20 agronomic traits and leaf phenotypic traits were observed and described. The data were used for phenotypic diversity analysis, correlation analysis, principal component analysis and cluster analysis. The result showed that the genetic diversity index of 20 traits ranged from 0.82 to 4.37, among which the largest was the base depth and the smallest was the leaf width; the coefficient of variation of the 12 quantitative traits ranged from 10.55% to 41.47%. the highest coefficient of variation was the height of dead leaves, and the smallest was the content of chlorophyll, except for the angle of branches, all the quantitative characters tended to be normal distribution. The correlation analysis showed that 28 pairs of traits had significant correlation(P<0.01), and 13 pairs had significant correlation(P<0.05). According to principal component analysis, 20 traits were simplified into 9 principal components, and the cumulative contribution rate was 73.414%, nine traits including plant height, dead leaves heigh, stem diameter, symmetry of leave base, stipule, leaf tip shape, depth of the first pair of lobes, depth of the second pair of lobes and leaf yield were selected as key indexes for evaluating agronomic traits and leaf phenotypic traits of A.argyi germplasm resources. With cluster analysis, 100 accessions of A.argyi were classified into 3 groups, the groupⅠincluded the dwarf plants with thick stem and large leaf, the groupⅡincluded high plants with wide leaf and high yield, the group Ⅲ included dwarf plants with thin stem and flat bottom shape of leaf, which could provide the basis for cultivation identification and variety breeding of A.argyi germplasm resources.
Artemisia ; China ; Phenotype ; Plant Breeding ; Plant Leaves/genetics*

As a natural plant source of artemisinin,a first-line drug against malaria,Artemisia annua directly affects the extraction process of artemisinin and the source of artemisinin. At present,traditional breeding methods combined with tissue culture are often used to breed high-yield artemisinin-containing new varieties of A. annua. However,the breeding method has the disadvantages of low efficiency and continuous selection. In this study,heavy ion beam irradiation technology was used to observe the specific germplasm resources of A. annua,and the morphological characteristics,agronomic traits and artemisinin content were used as indicators to observe the selection materials and materials. The cultivated new varieties were compared with trials and regional trials. In addition,the new variety of A. annua was identified by SRAP molecular marker technology. The results showed that the new variety of A. annua, " Kehao No.1",had an average yield of 235. 0 kg of dry leaf per mu,which was more than 20% higher than that of the control. Especially,the average artemisinin content was 2. 0%,which was 45% higher than that of the control,and the " Kehao No.1" has high anti-white powder disease,high-yield and high-quality new varieties. Therefore,mutagenic breeding of heavy ion beam irradiation can significantly improve the yield and artemisinin content of the " Kehao No. 1" and it has a good promotion value.
Artemisia annua/genetics* ; Artemisinins/analysis* ; Heavy Ions ; Mutagenesis ; Phenotype ; Plant Breeding ; Plants, Medicinal/genetics*