Complete chloroplast genome of Ligustrum lucidum and highly variable marker identification for Ligustrum / 中国中药杂志

Ligustri Lucidi Fructus, the sun-dried mature fruit of Ligustrum lucidum, is cool, plain, sweet, and bitter, which can be used as both food and medicine, with the effects of improving vision, blacking hair, and tonifying liver and kidney. It takes effect slowly. However, little is known about the genetic information of the medicinal plant and it is still a challenge to distinguish Ligustrum species. In this study, the complete chloroplast genome of L. lucidum was obtained by genome skimming and then compared with that of five other Ligustrum species, which had been reported. This study aims to evaluate the interspecific variation of chloroplast genome within the genus and develop molecular markers for species identification of the genus. The result showed that the chloroplast genome of L. lucidum was 162 162 bp with a circular quadripartite structure of two single-copy regions separated by a pair of inverted repeats. The Ligustrum chloroplast genomes were conserved with small interspecific difference. Comparative analysis of six Ligustrum chloroplast genomes revealed three variable regions(rbcL-accD, ycf1a, and ycf1b), and ycf1a and ycf1b can be used as the species-specific DNA barcode for Ligustrum. Phylogeny analysis provided the best resolution of Ligustrum and supported that L. lucidum was sister to L. gracile. This study clarified the genetic diversity of L. lucidum from provenance, which can serve as a reference for further analysis of pharmacological differences and breeding of excellent varieties with stable drug effects.

Molecular identification and alkaloids content of endangered Chinese medicinal plant Coptis chinensis var. brevisepala / 中国中药杂志

According to the records of Chinese materia medica,Coptis chinensis var. brevisepala is an authentic Chinese medicinal plant highly recommended by ancient physicians since its rhizome is like a string of beads and has a good medicinal value. However,its medicinal components and values remain to be studied as it is endangered because of overexploitation. Therefore,this study aims to quantitatively determine its effective components based on UPLC-QTOF-MS,and to compare the contents of isoquinoline alkaloids in C.chinensis var. brevisepala with those in other Coptis species. Meanwhile,molecular methods accurately identified 12 batches of C. chinensis var. brevisepala,9 batches of C. chinensis,4 batches of C. deltoidea,and 1 batch of C. teeta. Gradient elution was performed with Waters CORTECS C18 column( 4. 6 mm× 150 mm,2. 7 μm) and the mobile phase acetonitrile-water with 0. 4% formic acid. Mass spectrometry was conducted in ESI positive mode. The quantitative results showed that 8 main alkaloids had a good linear relationship within the concentration range( R~2>0. 996),with the recovery rate of 95. 18%-105. 0% and the RSD of 0. 28%-3. 7%. Compared with that of other Coptis species,the rhizome of C. chinensis var. brevisepala had the highest contents of berberine and columbamine. The total content of the 8 alkaloids in C. chinensis var. brevisepala was similar to that in C. chinensis but higher than that of the other two species. PCA was performed to compare the alkaloids among the 4 species. Besides,the 8 alkaloids were evaluated in different parts of C. chinensis var. brevisepala. The results indicate that this method is reliable and efficient and can provide a reference for the quality research.

Research progress on medicinal resources of Coptis and its isoquinoline alkaloids / 中国中药杂志

Coptidis Rhizoma is a common Chinese medicinal in clinical practice,with the effects of clearing heat,drying dampness,purging fire,and removing toxin. All the medicinal plants of Coptis can be used for clinical treatment,but some species are endangered due to resource destruction and difficulty in planting. The dominant medicinal components in Coptidis Rhizoma are isoquinoline alkaloids. There are various methods for the analysis and detection of alkaloids,such as LC-MS,HPLC,and TLC,among which LC-MS is the most widely applied. Different plants of Coptis vary in the kind and content of alkaloids. C. chinensis,C. deltoidea,C. teeta,C. chinensis var. brevisepala,C. omeiensis,C. quinquefolia,and C. quinquesecta mainly contain berberine,palmatine,coptisine,jatrorrhizine,and columbamine,five effective alkaloid components. Plant isoquinoline alkaloids( PIAs) have strong pharmacological activity but are difficult to prepare. The application of synthetic biology of PIAs will be helpful for the clinical application of PIAs. This paper reviews the research progress on biological resources of Coptis species and structures of alkaloids as well as analysis methods and synthetic biology for isoquinoline alkaloids in the medicinal plants of Coptis in recent years,which will facilitate the protection of Coptis medicinal resources and the application and development of alkaloids.

Effects and mechanisms of nitrogen application on stress resistance of Chinese materia medica / 中国中药杂志

Nitrogen fertilizers play an important role in the regulation of plant stress resistance. Impacts of nitrogen fertilizers on abiotic stress resistance and biotic stress resistance of Chinese materia medica(CMM) were summarized in this study. Adequate nitrogen application improves the abiotic stress resistance and weed resistance of CMM, however adverse effect appears when excess nitrogen is used. Generally, pest resistance decreases along with nitrogen deposition, while effects of nitrogen application on disease resistance vary with different diseases. Mechanisms underlying the impact of nitrogen fertilizers on plant stress resistance were also elucidated in this study from three aspects including physical defense mechanisms, biochemistry mechanisms and molecular defense mechanisms. Nitrogen availability modulates physical barrier of CMM like plant growth, formation of lignin and wax cuticle, and density of stomata. Growth of CMM promoted by nitrogen fertilizer may cause some decrease in pest resistance of CMM due to an increase in hiding places for pest along with plant growth. High ambient humidity caused by excessive plant growth facilitates the growth and development of CMM pathogen. Nitrogen application can also interfere with the accumulation of lignin in CMM which makes CMM more vulnerable to pest and pathogen attack. Stomatal closing delays due to nitrogen application is also a causal factor of increasing pathogen infection after nitrogen deposition. Biochemical defenses of plants are mainly achieved through nutrient elements, secondary metabolites, defense-related enzymes and proteins. Nutritional level of CMM and various antioxidant enzymes and resistance-related protein activities are elevated along with nitrogen deposition. These antioxidant enzymes can reduce the damage of reactive oxygen species content produced by plant in response to adversity and therefore enhance stress resistance of CMM. Researches showed that nitrogen application could also cause an increase in nitrogen-containing secondary metabolites content and a decrease in non-nitrogen-containing secondary metabolites content respectively. Nitrogen-mediated molecular defense mechanisms includes multiple plant hormones and nitric oxide signals. Plant hormones related to plant defense like salicylic acid, jasmonic acid and abscisic acid can be modulated by nitrogen application. Negative effect of nitrogen deposition was found on salicylic acid accumulation and the expression of related plant disease resistance genes. However, jasmonic acid level can be elevated by nitrogen. Nitric oxide signals constitute an important part of nitrogen mediated defense mechanisms. Nitric oxide signaling is related to many aspects of plant immunity. The roles of nitrogen fertilizers in CMM stress resistance are complex and may vary with different CMM varieties and environments. Further studies are urgently needed to provide a comprehensive understanding of how to improve stress resistance of CMM by using fertilizers.

Current situation of nitrogen application and its effects on yield and quality of Chinese materia medica / 中国中药杂志

Nitrogen fertilizer has been the long-lasting crucial component in cultivation of Chinese materia medica(CMM) and crops for its profound effects on enhancing the productivity. In consideration of its role in better production, intensive and excessive application of N fertilizer is often found in CMM cultivation. Therefore, firstly, this review summarized various concentrations of N application with regards to different CMM and districts from the literatures published in the last two decades. The recommended concentration of nitrogen application of forty seven CMM species were covered in this review. We found that the optimum rates of nitrogen fertili-zer for different medicinal plants species were varied in the range between 0-1 035.55 kg·hm~(-2). Most of the optimum rates of nitrogen fertilizer for CMM in published researches fell between 100-199 kg·hm~(-2). The optimum rate of nitrogen fertilizer is not only related to amount of nitrogen required for different medicinal plants but also to soil fertilities of different fields. In addition, we outlined the diffe-rent effects of proper and excessive nitrogen deposition on yield of CMM. Proper nitrogen deposition benefits the yield of CMM, howe-ver, excessive nitrogen use accounts for a decrease in CMM yield. We elucidated that nutritional content, water use efficiencies, and photosynthesis capacity were major influencing factors. Researches showed that proper nitrogen fertilizer could promote the water use efficiencies of plants and boost photosynthesis. Consequently, the yield of CMM can be enhanced after nitrogen deposition. However, negative effects of nitrogen fertilizer were also found on plant including producing toxic substances to the soil and causing severe pest damages. Lastly we analyzed the impact of N fertilizer application on secondary metabolites which accounts for a large part of active pharmaceutical ingredients of CMM. It usually caused an increase in nitrogen-containing secondary metabolites content and a decrease in non-nitrogen-containing secondary metabolites content respectively. The potential underlying mechanisms are the different synthetic pathways of these metabolites and the plant nutritional status. Synthesis of non-nitrogen-containing secondary metabolites like phenols can be inhibited after nitrogen application because of the competition of the same precursor substances between metabolites synthesis and plant growth. To sum up, impacts and mechanisms of nitrogen fertilizer on yield and quality enhancement of CMM were discussed in this review. Negative effects of excessive nitrogen application on CMM should be paid special attention in CMM cultivation and prescription fertilization based on the field soil quality is strongly recommended. Overall, this review aims to provides insights on improving the proper application of N fertilizer in the cultivation of CMM.

Biological characteristics of wild Angelica sinensis seed / 中国中药杂志

Wild Angelica sinensis is almost endangered, studying the biological characteristics of wild A. sinensis seeds is helpful for varietal improvement and its conservation. This paper systematically studied the morphological structure, thousand seed weight, viability, storage and other basic biological characteristics of wild A. sinensis fruits and seeds, and focused on the germination of excised embryos,the development of embryo, the effects of the temperature,light and hormones on seed germination.The study found that:①The embryos are not fully developed when harvested, the initial germination rate was low, the embryos can develop. After 15 days of low temperature storage, the embryos can develop completely and the germination rate is significantly increased. These results show that wild A. sinensis seeds have no dormancy, and the low germination rate is due to the low maturity of wild A. sinensis seeds. ②The sui-table germination temperature of wild A. sinensis is 15-25 ℃,and the optimal temperature is 20 ℃. Light does not affect the germination of A. sinensis seeds.③The applicable concentration of GA_3 can promote seeds germination, IAA and 6-BA has no significant effect on germination.④The optimum storage condition is dry storage at 4 ℃. Wild A. sinensis seeds can be stored for 1.5 years and cultivated seeds can be stored for 1 year.During the introduction and conservation, the best treatment conditions were dry storage at 4 ℃ for 30 d and soaking seeds with 200 mg·L~(-1) GA_3, the germination rate can reach 86.7%.

Population characteristics and threatened of wild Angelica sinensis in Gansu province / 中国中药杂志

The study aims to investigate spatial distribution pattern and age structure of wild Angelica sinensis in Gansu province.Ten plots each with an area of 100 m2 were set and the spatial coordinates of all wild A. sinensis individuals were measured within each plot. Based on plant individual mapping data,we explored the spatial distribution pattern and its differences between different life history stages of wild A. sinensis in Gansu province by using nearest neighbor distance statistics. Correlation analysis were carried out to explore the relationship between spatial aggregation degree and topographic factors. We also distinguished individuals to three life history stages( i.e. seedlings,adults and boltings) and then test the differences among/between them using nonparametric test.(1)We found that the dominant spatial distribution pattern of wild A. sinensis population in Gansu was aggregated distribution. There was no significant correlation between spatial aggregation degree of wild A. sinensis and altitude,slope and aspect. There was no significant difference between the average distance from seedlings to their nearest bolting individuals; the average distance from adult individuals to their nearest seedlings was significantly larger than the average distance from adult individuals to their nearest adult individuals; and the average distance from bolting individuals to their nearest adult individuals was significantly smaller than the average distance from bolting individuals to their nearest bolting individuals.(2)The age structure was showed as a declining population,characterized by less seedlings and bolting individuals,while more adult individuals within population. The population characteristics of wild A. sinensis,characterized by aggregated distribution pattern and senescent type of age structure,are disadvantage to its population development. The factors,such as habitat specialization,human activities and intraspecific competition,which shapes the current population characteristics,may increase the threatened status of wild A. sinensis. We suggest to strengthen the protection of wild A. sinensis.

Cloning and prokaryotic expression analysis of HDS from Salvia miltiorrhiza bge.f.alba / 药学学报

According to the designed specific primers of gene fragment based on the Salvia miltiorrhiza transcriptome data, with the method of reverse transcription polymerase chain reaction (RT-PCR), this study cloned full-length cDNA sequence of 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate synthase gene from Salvia miltiorrhiza bge.f.alba, this sequence is named as SmHDS and its GenBank registration number is KJ746807. SmHDS, 2 529 bp long, contains an ORF of 2 229 bp, encodes 742 amino acids, including 5' UTR 170 bp and 3' UTR 130 bp. Using bioinformatics software, having made a homology analysis of the obtained sequence, we can have a conclusion that SmHDS have a close genetic relationship with HDS of Salvia miltiorrhiza. Analysis result of prokaryotic expression revealed that in Escherichia coli, SmHDS expressed target proteins which in size are comparable with the protein predicted. Meanwhile, the 4 factors which can influence the protein expression were optimized, the 4 factors are inducing temperature, inducing time, IPTG concentrations and density of inducing host bacterium (A600). The optimal expression conditions of SmHDS were 30 degrees C until the A600 is 0.6, and add IPTG to a final concentration of 0.2 mmol x L(-1), and the induction time of 20 h. It provides theoretical basis for the further study of the function of 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate synthase in the biosynthesis of tanshinone compounds.

Key problems in development of molecular identification in traditional Chinese medicine / 中国中药杂志

Molecular identification of Chinese traditional medicine has come from laboratory research into application, but there are some misunderstandings and problems emerging after rapid development. In this paper, we discuss the usage principle, hot field and technology innovation in molecular identification of Chinese traditional medicine. And molecular identification of traditional Chinese medicine has scientific and objective basis, follows the certain systematic research background, and adopts practical principles to establish case by case multi-class identification system. In order to achieve rapid, on-site, high throughput, low cost of traditional Chinese medicine identification purpose, molecular identification technology is further developing for meet the actual needs and the laboratory results further transformation in the service of traditional Chinese medicine industry.

Prospect and application of microsatellite population genetics in study of geoherbs / 中国中药杂志

The author introduces the basic concepts of microsatellite and population genetics and its characteristics, expounds the application of these theories for population genetic structure and genetic diversity, gene flow and evolutionary significant unit ESU division research. This paper discuss its applicationin study of genetic causes, origin of cultivation, different regional origins of geoherbs, aiming at providing a new theory and method for geoherbs.

Molecular identification of astragali radix and its adulterants by ITS sequences / 中国中药杂志

<p><b>OBJECTIVE</b>To explore a new method for identification Astragali Radix from its adulterants by using ITS sequence.</p><p><b>METHOD</b>Thirteen samples of the different Astragali Radix materials and 6 samples of the adulterants of the roots of Hedysarum polybotrys, Medicago sativa and Althaea rosea were collected. ITS sequence was amplified by PCR and sequenced unidirectionally. The interspecific K-2-P distances of Astragali Radix and its adulterants were calculated, and NJ tree and UPGMA tree were constructed by MEGA 4.</p><p><b>RESULT</b>ITS sequences were obtained from 19 samples respectively, there were Astragali Radix 646-650 bp, H. polybotrys 664 bp, Medicago sativa 659 bp, Althaea rosea 728 bp, which were registered in the GenBank. Phylogeny trees reconstruction using NJ and UPGMA analysis based on ITS nucleotide sequences can effectively distinguish Astragali Radix from adulterants.</p><p><b>CONCLUSION</b>ITS sequence can be used to identify Astragali Radix from its adulterants successfully and is an efficient molecular marker for authentication of Astragali Radix and its adulterants.</p>

A taxonomic study on the original plant of radix astragali / 药学学报

There are dispute about the status of taxonomy among Astragalus membranaceus (Fisch.) Bge, A. membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao. and A. pallidipurpureus stat. nov. The varieties and taxa of the complex are still in need of revision. With molecular biology study used trnH-psbA intergenic region, the taxonomic revision of Radix Astragali has been made. A. pallidipurpureus stat. nov is suggested as a new species.

Genetic relationships of Atractylodes plants / 药学学报

The phylogenetic relationships of the genus Atractylodes DC. was analyzed using the nuclear ribosomal ITS and three chloroplast fragments, including atpB-rbcL, psbB-psbF and trnL-trnF intergenic spacer (IGS) sequences. Phylogenetic analyses revealed that A. lancea subsp. luotianensis and A. lancea, A. chinensis var. liaotungensis and A. coreana form monophyletic terminal clade, separately. The trees, within each the pairwise genetic distances, did not support Hu's classification. Authors suggested that A. lancea Subsp. luotianensis should be included in A. lancea rather than be treated as a separate subspecies. A. carlinoides was placed in the basal position of Atractylodes, which had a distant relationship with the others of the genus. The results lead us to suggest that A. chinensis var. liaotungensis be put into A. coreana, A. chinensis as a subspecies of A. lancea.

RAPD analysis on genetic relationship among varieties of Rehmannia glutinosa / 中国中药杂志

<p><b>OBJECTIVE</b>To define the genetic relationship and other genetic characters of different varieties of Rehmannia glutinosa.</p><p><b>METHOD</b>RAPD reactions were conducted on total 54 samples from 16 varieties and 2 populations of R. glutinosa. The data were analyzed by Phylips, Popgene and Arlequin software.</p><p><b>RESULT</b>The majority rule consensus tree defined the 54 samples to 3 groups. Population genetics analysis and AMOVA showed that the genetic diversity among varieties/populations much greater than that within varieties/populations, but within group II the genetic diversity among varieties was similar to that within varieties.</p><p><b>CONCLUSION</b>Due to the long-time asexual propagation of R. glutinosa, some genetic differentiation has been accumulated and fixed within the species. It was shown that the genetic distance between wild population and cultivated varieties was greater than the genetic distance among cultivated varieties. The wild resource of the plant should be paid more attention and studies.</p>