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Abstract Background The sustainable supply of medicinal plants is important, and cultivating and domesticating them has been suggested as an optimal strategy. However, this can lead to a loss of genetic diversity. Tripterygium wilfordii Hook. f. is a medicinal plant commonly used in traditional Chinese medicine, but its wild populations are dwindling due to excessive harvesting. To protect the species and meet the increasing demand, it is urgent to cultivate it on a large scale. However, distinguishing between T. wilfordii and T. hypoglaucum, two similar species with different medicinal properties, is challenging. Therefore, it is crucial to understand the genetic diversity and population structure of these species for their sustainable utilization. Results In this study, we investigated the genetic diversity and population structure of the two traditional medicinal semiwoody vines plant species, Tripterygium wilfordii and T. hypoglaucum, including wild and cultivated populations using chloroplast DNA (cpDNA) sequences and microsatellite loci. Our results indicated that the two species maintain a high level of genetic divergence, indicating possible genetic bases for the different contents of bioactive compounds of the two species. T. wilfordii showed lower genetic diversity and less subdivided population structures of both markers than T. hypoglaucum. The potential factors in shaping these interesting differences might be differentiated pollen-to-seed migration rates, interbreeding, and history of population divergence. Analyses of cpDNA and microsatellite loci supported that the two species are genetically distinct entities. In addition, a significant reduction of genetic diversity was observed for cultivated populations of the two species, which mainly resulted from the small initial population size and propagated vegetative practice during their cultivation. Conclusion Our findings indicate significant genetic divergence between T. wilfordii and T. hypoglaucum. The genetic diversity and population structure analyses provide important insights into the sustainable cultivation and utilization of these medicinal plants. Accurate identification and conservation efforts are necessary for both species to ensure the safety and effectiveness of crude drug use. Our study also highlighted the importance of combined analyses of different DNA markers in addressing population genetics of medicinal plants because of the contrasts of inheritance and rates of gene flow. Large-scale cultivation programs should consider preserving genetic diversity to enhance the long-term sustainability of T. wilfordii and T. hypoglaucum. Our study proposed that some populations showed higher genetic diversity and distinctness, which can be considered with priority for conservation and as the sources for future breeding and genetic improvement.

Abstract Background Leonurus japonicus, a significant medicinal plant known for its therapeutic effects on gynecological and cardiovascular diseases, has genetic diversity that forms the basis for germplasm preservation and utilization in medicine. Despite its economic value, limited research has focused on its genetic diversity and divergence. Results The avg. nucleotide diversity of 59 accessions from China were 0.00029 and hotspot regions in petN-psbM and rpl32-trnL (UAG) spacers, which can be used for genotype discrimination. These accessions divided into four clades with significant divergence. The four subclades, which split at approximately 7.36 Ma, were likely influenced by the Hengduan Mountains uplift and global temperature drop. The initial divergence gave rise to Clade D, with a crown age estimated at 4.27 Ma, followed by Clade C, with a crown age estimated at 3.39 Ma. The four clades were not showed a clear spatial distribution. Suitable climatic conditions for the species were identified, including warmest quarter precipitation 433.20 mm ~ 1,524.07 mm, driest month precipitation > 12.06 mm, and coldest month min temp > -4.34 °C. The high suitability distribution showed contraction in LIG to LGM, followed by expansion from LGM to present. The Hengduan Mountains acted as a glacial refuge for the species during climate changes. Conclusions Our findings reflected a clear phylogenetic relationships and divergence within species L. japonicus and the identified hotspot regions could facilitate the genotype discrimination. The divergence time estimation and suitable area simulation revealed evolution dynamics of this species and may propose conservation suggestions and exploitation approaches in the future.

Angelica sinensis (Oliv.) Diels belongs to the Apiaceae family. The root of A. sinensis, is used in traditional Chinese medicine for its antioxidant and immune regulation properties. The main active compounds in A. sinensis include organic acids, phthalides and coumarins, and their biosynthetic pathways are the focus of international attention. A. sinensis is prone to early flowering and bolting, which negatively impacts production for several reasons, including germplasm degradation and quality instability in artificial cultivation. The identification of top-geoherbalism of A. sinensis has also become the focus of recent research, as it would allow selection for breeds with excellent medicinal quality and remarkable curative effects. Advances in sequencing technology and bioinformatic methodologies have enabled extensive molecular and genetic studies in A. sinensis. In this review, we summarize the latest molecular research advances related to A. sinensis, including biosynthetic pathways and regulation of active compounds, and molecular underpinnings of early bolting and flowering and top-geoherbalism. We discuss limitations of the current research and propose prospective topics in need of further exploration.

Coptis huanjiangensis, a new species of Ranunculaceae distributed in the valleys of Jiuwanshan National Natural Reserve in Huanjiang county (Guangxi, China), is described and illustrated for the first time based on morphological and plastome sequences data. It differs from C. chinensis, C. deltoidei and C. omeiensis mainly by having notably longer petiole, scape, bigger leaf blade with lobes obviously remote and robust rhizomes without stolons. Phylogenetic analyses support that C. huanjiangensis is sister to C. omeiensis and C. deltoidei.

The genus Coptis belongs to the Ranunculaceae family, containing 15 recognized species highly diverse in morphology. It is a conspicuous taxon with special evolutionary position, distribution pattern and medicinal value, which makes it to be of great research and conservation significance. In order to better understand the evolutionary dynamics of Coptis and promote more practical conservation measures, we performed plastome sequencing and used the sequencing data in combination with worldwide occurrence data of Coptis to estimate genetic diversity and divergence times, rebuild biogeographic history and predict its potential suitable distribution area. The average nucleotide diversity of Coptis was 0.0067 and the hotspot regions with the highest hypermutation levels were located in the ycf1 gene. Coptis is most likely to have originated in North America and Japanese archipelago and has a typical Eastern Asian and North American disjunct distribution pattern, while the species diversity center is located in Mid-West China and Japan. The crown age of the genus is estimated at around 8.49 Mya. The most suitable climatic conditions for Coptis were as follows: precipitation of driest quarter > 25.5 mm, annual precipitation > 844.9 mm and annual mean temperature -3.1 to 19 °C. The global and China suitable area shows an upward trend in the future when emission of greenhouse gases is well controlled, but the area, especially in China, decreases significantly without greenhouse gas policy interventions. The results of this study provide a comprehensive insight into the Coptis evolutionary dynamics and will facilitate future conservation efforts.

Abstract Background Most Distylium species are endangered. Distylium species mostly display homoplasy in their flowers and fruits, and are classified primarily based on leaf morphology. However, leaf size, shape, and serration vary tremendously making it difficult to use those characters to identify most species and a significant challenge to address the taxonomy of Distylium. To infer robust relationships and develop variable markers to identify Distylium species, we sequenced most of the Distylium species chloroplast genomes. Results The Distylium chloroplast genome size was 159,041–159,127 bp and encoded 80 protein-coding, 30 transfer RNAs, and 4 ribosomal RNA genes. There was a conserved gene order and a typical quadripartite structure. Phylogenomic analysis based on whole chloroplast genome sequences yielded a highly resolved phylogenetic tree and formed a monophyletic group containing four Distylium clades. A dating analysis suggested that Distylium originated in the Oligocene (34.39 Ma) and diversified within approximately 1 Ma. The evidence shows that Distylium is a rapidly radiating group. Four highly variable markers, matK-trnK, ndhC-trnV, ycf1, and trnT-trnL, and 74 polymorphic simple sequence repeats were discovered in the Distylium plastomes. Conclusions The plastome sequences had sufficient polymorphic information to resolve phylogenetic relationships and identify Distylium species accurately.

Root rot has been a major problem for cultivated populations of Coptis chinensis var. chinensis in recent years. C. chinensis var. brevisepala, the closest wild relative of C. chinensis var. chinensis, has a scattered distribution across southwestern China and is an important wild resource. Genetic diversity is associated with greater evolutionary potential and resilience of species or populations and is important for the breeding and conservation of species. Here, we conducted multiplexed massively parallel sequencing of the plastomes of 227 accessions of wild and cultivated C. chinensis using 111 marker pairs to study patterns of genetic diversity, population structure, and phylogeography among wild and cultivated C. chinensis populations. Wild and cultivated resources diverged approximately 2.83 Mya. The cultivated resources experienced a severe genetic bottleneck and possess highly mixed germplasm. However, high genetic diversity has been retained in the wild resources, and subpopulations in different locations differed in genotype composition. The significant divergence in the genetic diversity of wild and cultivated resources indicates that they require different conservation strategies. Wild resources require in situ conservation strategies aiming to expand population sizes while maintaining levels of genetic diversity; by contrast, germplasm resource nurseries with genotypes of cultivated resources and planned distribution measures are needed for the conservation of cultivated resources to prevent cultivated populations from undergoing severe genetic bottlenecks. The results of this study provide comprehensive insights into the genetic diversity, population structure, and phylogeography of C. chinensis and will facilitate future breeding and conservation efforts.

Abstract Background Atractylodes DC is the basic original plant of the widely used herbal medicines “Baizhu” and “Cangzhu” and an endemic genus in East Asia. Species within the genus have minor morphological differences, and the universal DNA barcodes cannot clearly distinguish the systemic relationship or identify the species of the genus. In order to solve these question, we sequenced the chloroplast genomes of all species of Atractylodes using high-throughput sequencing. Results The results indicate that the chloroplast genome of Atractylodes has a typical quadripartite structure and ranges from 152,294 bp (A. carlinoides) to 153,261 bp (A. macrocephala) in size. The genome of all species contains 113 genes, including 79 protein-coding genes, 30 transfer RNA genes and four ribosomal RNA genes. Four hotspots, rpl22-rps19-rpl2, psbM-trnD, trnR-trnT (GGU) , and trnT (UGU) -trnL, and a total of 42–47 simple sequence repeats (SSR) were identified as the most promising potentially variable makers for species delimitation and population genetic studies. Phylogenetic analyses of the whole chloroplast genomes indicate that Atractylodes is a clade within the tribe Cynareae; Atractylodes species form a monophyly that clearly reflects the relationship within the genus. Conclusions Our study included investigations of the sequences and structural genomic variations, phylogenetics and mutation dynamics of Atractylodes chloroplast genomes and will facilitate future studies in population genetics, taxonomy and species identification.

Abstract Background Angelica L. (family Apiaceae) is an economically important genus comprising ca. One hundred ten species. Angelica species are found on all continents of the Northern Hemisphere, and East Asia hosts the highest number of species. Morphological characters such as fruit anatomy, leaf morphology and subterranean structures of Angelica species show extreme diversity. Consequently, the taxonomic classification of Angelica species is complex and remains controversial, as the classifications proposed by previous studies based on morphological data and molecular data are highly discordant. In addition, the phylogenetic relationships of major clades in the Angelica group, particularly in the Angelica s. s. clade, remain unclear. Chloroplast (cp) genome sequences have been widely used in phylogenetic studies and for evaluating genetic diversity. Results In this study, we sequenced and assembled 28 complete cp genomes from 22 species, two varieties and two cultivars of Angelica. Combined with 36 available cp genomes in GenBank from representative clades of the subfamily Apioideae, the characteristics and evolutionary patterns of Angelica cp genomes were studied, and the phylogenetic relationships of Angelica species were resolved. The Angelica cp genomes had the typical quadripartite structure including a pair of inverted repeats (IRs: 5836–34,706 bp) separated by a large single-copy region (LSC: 76,657–103,161 bp) and a small single-copy region (SSC: 17,433–21,794 bp). Extensive expansion and contraction of the IR region were observed among cp genomes of Angelica species, and the pattern of the diversification of cp genomes showed high consistency with the phylogenetic placement of Angelica species. Species of Angelica were grouped into two major clades, with most species grouped in the Angelica group and A. omeiensis and A. sinensis grouped in the Sinodielsia with Ligusticum tenuissimum. Conclusions Our results further demonstrate the power of plastid phylogenomics in enhancing the phylogenetic reconstructions of complex genera and provide new insights into plastome evolution across Angelica L.

Bactericera gobica is the major pest of Goji berry plants and causes severe damage. Psyllids mainly use the antennal sensilla to recognize olfactory cues necessary to find host plants and mates. However, the structure and function of the antenna and the antennal sensilla of B. gobica remains previously unexplored. Here, we identify the external and internal morphology of the antennal sensilla of B. gobica using both scanning electron microscopy (SEM) and transmission electron microscopy (TEM). We found seven types of sensilla on the filiform antennae, including apical setae (LAS, SAS), sensilla basiconica (SB1, SB2), sensilla campaniform (SCA), sensilla chaetica (ChS1, ChS2), cavity sensilla (CvS1, CvS2), antennal rhinaria (AR1, AR2), and sensilla trichodea (ST). Five of these sensilla types—apical setae, sensilla basiconica, sensilla chaetica, cavity sensilla, and antennal rhinaria—may have olfactory functions based on their porous surfaces and internal dendritic outer segments (DOS). We also found several differences between the two sexes of B. gobica in the sensilla array and internal structure. ChS and DOS in the protrusions of AR were more abundant in males than females. Altogether, we comprehensively revealed the fine structure and probable function of B. gobica antennae and identified differences in the distribution and structure between psyllid sexes. Our findings provide important insights for future studies on defining the olfactory function of psyllid antenna using electrophysiological methods.

Leonurus cardiaca has a long history of use in western herbal medicine and is applied for the treatment of gynaecological conditions, anxiety, and heart diseases. Because of its botanical relationship to the primary Chinese species, L. japonicus, and extensive medical indications that go beyond the traditional indications for the Chinese species, it is a promising medicinal resource. Therefore, the features of genetic diversity and variability in the species have been prioritized. To explore these issues, we sequenced the chloroplast genomes of 22 accessions of L. cardiaca from different geographical locations worldwide using high-throughput sequencing. The results indicate that L. cardiaca has a typical quadripartite structure and range from 1,51,236 bp to 1,51,831 bp in size, forming eight haplotypes. The genomes all contain 114 distinct genes, including 80 protein-coding genes, 30 transfer RNA genes and four ribosomal RNA genes. Comparative analysis showed abundant diversity of single nucleotide polymorphisms (SNPs), indels, simple sequence repeats (SSRs) in 22 accessions. Codon usage showed highly similar results for L. cardiaca species. The phylogenetic and network analysis indicated 22 accessions forming four clades that were partly related to the geographical distribution. In summary, our study highlights the advantage of chloroplast genome with large data sets in intraspecific diversity evaluation and provides a new tool to facilitate medicinal plant conservation and domestication.

Prunus japonica is an ornamental and medicinal plant that is widely cultivated. The complete chloroplast genome of P. japonica was sequenced using Illumina Hiseq X Ten platform. The chloroplast genome was 158,080 bp in length, containing two short inverted repeat (IRa and IRb) regions of 26,385 bp, which was separated by a large single copy (LSC) region of 86,270 bp and a small single copy (SSC) region of 19,040 bp. The GC content of the whole chloroplast genome was 36.8%. The chloroplast DNA of P. japonica comprised 112 distinct genes, including 78 protein-coding genes, 4 ribosomal RNA genes and 30 transfer RNA genes. Phylogenetic analysis indicated that all species of Prunus formed a monophyletic group, P. japonica was closely related to P. hulimis.

Sargentodoxa cuneata is used as traditional Chinese medicine. In this study, we report its complete chloroplast genome by Illumina pair-end sequencing. The total chloroplast (cp) genome size was 158,094 bp in length, containing a pair of inverted repeats of 26,132 bp, separated by large single-copy and small single-copy regions of 86,508 bp and 19,322 bp, respectively. The chloroplast genome of S. cuneata encodes 113 different genes, including 79 protein-coding genes, 30 transfer RNAs, and 4 ribosomal RNAs. A total of 84 perfect chloroplast microsatellites were analyzed in the S. cuneata. The majority of the SSRs in this chloroplast genome are mononucleotides (66.67%). The reconstructed phylogeny revealed that S. cuneata was sister to the remaining Lardizabalaceae.

Crataegus kansuensis Wils. is an important wild eco economical species of the family Rosaceae. The complete chloroplast genome reported here is 159,865 bp in length, including two inverted repeats (IRs) of 26,384 bp, which are separated by a large single-copy (LSC) and a small single-copy (SSC) of 87,815 bp and 19,282 bp, respectively. The whole chloroplast genome of C. kansuensis contains 113 genes, including 79 protein-coding genes, 30 transfer RNA, and 4 ribosome RNA. Phylogenetic analysis indicated that C. kansuensis is closely related to that of C. chungtienensis and C. marshallii, and the genus Crataegus L. was sister to the genus Amelanchier Medik.

Abutilon theophrasti Medik. is an annual weed, widely distributed in Asia and Europe. The complete chloroplast genome reported here is 160,446 bp in length, including two inverted repeats (IRs) of 25,064 bp, which are separated by a large single-copy (LSC) and a small single-copy (SSC) of 89,089 and 21,229 bp, respectively. The whole chloroplast genome of A. theophrasti contains 113 distinct genes, including 79 protein-coding genes, 30 transfer RNA, and four ribosome RNA. Phylogenetic analysis indicated that A. theophrasti is located in the basal position in Malveae.

Mume Fructus (MF) contains a variety of organic acids, free amino acids, and nucleoside components, and studies have not yet analyzed the relationship between the components of free amino acids and nucleosides with the varieties of MF. A rapid and sensitive method was established for simultaneous determination of 21 free amino acids and 9 nucleosides in MF by ultrafast liquid chromatography-mass spectrometry. The analysis was carried out on a Waters XBridge Amide column (100 mm × 2.1 mm, 3.5 μm) with elution by the mobile phase of 0.2% aqueous formic acid (A) and 0.2% formic acid acetonitrile (B) at a flow rate of 0.2 mL/min with 1 μL per injection. The column temperature was maintained at 30°C. The target compounds were analyzed by the positive ion multiple reaction monitoring (MRM) mode. The comprehensive evaluation of the samples was carried out by principal component analysis (PCA) and technique for order preference by similarity to an ideal solution (TOPSIS) analysis. Results showed the method could simultaneously determine 30 components in MF. The content of total analytes in six mainstream varieties was different, exhibited the order Nangao > Daqingmei > Zhaoshuimei > Yanmei > Shishengme > Baimei, and aspartic acid and adenosine were the most abundant amino acid and nucleoside. PCA and OPLS-DA could easily distinguish the samples, and 11 components could be chemical markers of sample classification. TOPSIS implied that the quality of Nangao and Daqingmei was superior to the other varieties. The results could provide a reliable basis for quality evaluation and utilisation of medicinal and edible MF.

Dioscorea L., the largest genus of the family Dioscoreaceae with over 600 species, is not only an important food but also a medicinal plant. The identification and classification of Dioscorea L. is a rather difficult task. In this study, we sequenced five Dioscorea chloroplast genomes, and analyzed with four other chloroplast genomes of Dioscorea species from GenBank. The Dioscorea chloroplast genomes displayed the typical quadripartite structure of angiosperms, which consisted of a pair of inverted repeats separated by a large single-copy region, and a small single-copy region. The location and distribution of repeat sequences and microsatellites were determined, and the rapidly evolving chloroplast genome regions (trnK-trnQ, trnS-trnG, trnC-petN, trnE-trnT, petG-trnW-trnP, ndhF, trnL-rpl32, and ycf1) were detected. Phylogenetic relationships of Dioscorea inferred from chloroplast genomes obtained high support even in shortest internodes. Thus, chloroplast genome sequences provide potential molecular markers and genomic resources for phylogeny and species identification.

Bupleurum chinense DC. is a traditional medicinal herb species widely distributed in most provinces of China. In this study, we constructed and annotated a complete circular chloroplast (cp) genome of B. chinense. The cp genome of B. chinense is 155,545 bp in length, including two inverted repeat (IR) regions of 26,305 bp, separated by a large single-copy (LSC) region of 85,430 bp and a small single-copy (SSC) region of 17,505 bp. The GC content of whole cp genome is 37.68%. The genome contains 113 different genes, including 79 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. A maximum-likelihood phylogenomic analysis showed that Bupleurum formed a monophyletic group, and was sister to other groups of Apiaceae.

In this study, we sequenced the complete chloroplast genome of Dracaena cochinchinensis and used the data to assess its genomic resources. The complete chloroplast genome revealed a circular genome of 155,182 bp size with 37.5% GC content. The chloroplast genome is a typical quadripartite chloroplast structure with inverted repeats (26,507 bp) separated by small single copy (18,466 bp) and large single copy (83,702 bp) regions. De novo assembly and annotation showed the presence of 112 unique genes with 78 protein-coding genes, 30 tRNA genes, and four rRNA genes. We identified a total of 68 simple sequence repeat (SSR) markers in the chloroplast genome. A maximum-likelihood phylogenomic analysis showed that the position of D. cochinchinensi was on the base of Nolinoideae. Overall, the results of this study will contribute to better support of the evolution, molecular biology and conservation of D. cochinchinensi.

Salvia miltiorrhiza Bunge，which is also known as a traditional Chinese herbal medicine，is widely studied for its ability to accumulate the diterpene quinone Tanshinones. In addition to producing a variety of diterpene quinone, S. miltiorrhiza Bunge also accumulates sterol, brassinosteroid and triterpenoids. During their biosynthesis, squalene synthase (SQS, EC 2.5.1.21) converts two molecules of the hydrophilic substrate farnesyl diphosphate into a hydrophobic product, squalene. In the present study, cloning and characterization of S. miltiorrhiza Bunge squalene synthase 2 (SmSQS2, Genbank Accession Number: KM408605) cDNA was investigated subsequently followed by its recombinant expression and preliminary enzyme activity. The full-length cDNA of SmSQS2 was 1 597 bp in length, with an open reading frame (ORF) of 1 245 bp encoding 414 amino acids. The deduced amino acid sequence of SmSQS2 shared high similarity with those of SQSs from other plants. To obtain soluble recombinant enzymes, the truncated SmSQS2 in which 28 amino acids were deleted from the carboxy terminus was expressed as GST-Tag fusion protein in Escherichia coli BL21 (DE3) and confirmed by SDS-PAGE and Western Blot analysis, and the resultant bacterial crude extract was incubated with farnesyl diphosphate and NADPH. GC-MS analysis showed that squalene was detected in the in vitro reaction mixture. The gene expression level was analyzed through Quantitative real-time PCR, and was found to be higher in roots as compared to the leaves, and was up-regulated upon YE+ Ag+ treatment. These results could serve as an important to understand the function of the SQS family. In addition, the identification of SmSQS2 is important for further studies of terpenoid and sterol biosynthesis in S. miltiorrhiza Bunge.