Genetic diversity analysis and fingerprints of Chrysanthemum×morifolium based on SSR molecular markers / 生物工程学报

The present study aims to explore the genetic diversity of germplasm resources of Chrysanthemum×morifolium (hereinafter, C.×morifolium) at the molecular level and to establish a fingerprint database of C.×morifolium varieties. We employed 12 pairs of primers with high levels of polymorphism, clear bands, and high degrees of reproducibility to analyze the SSR molecular markers and genetic diversity of 91 C.×morifolium materials and 14 chrysanthemum- related materials. With regard to constructing the fingerprints of the tested materials, we chose 9 pairs of core primers. The findings revealed that 12 primer pairs detected 104 alleles in 105 samples, ranging from 2 to 26. The average number of observed alleles (Na) per site was 9.25. The average number of effective alleles (Ne) per site was 2.745 6, with its range being 1.276 0 to 4.742 5. Shannon genetic diversity index (I) values ranged between 0.513 3 and 2.239 9 (M=1.209 0). Nei's gene diversity index (H) ranged between 0.216 3 and 0.789 1 (M=0.578 0). The observed heterozygosity (Ho) ranged between 0.223 3 and 0.895 2 (M=0.557 5). The expected heterozygosity (He) ranged between 0.217 4 and 0.793 3 (M=0.580 8). The polymorphism information content (PIC) ranged between 0.211 5 and 0.774 0 (M=0.532 9). The genetic similarity (GS) ranged between 0.228 5 and 1.000 0 (M=0.608 3). Cluster analysis revealed that when the genetic distance (GD) equals to 0.30, the tested materials can be classified into 2 groups. When the GD equals to 0.27, the first group can be divided into 6 subgroups; accordingly, 105 tested materials can be divided into 7 subgroups. The cophenetic correlation test was carried out based on the cluster analysis, and the corresponding results showed that the cluster map correlated with the genetic similarity coefficient (r=0.952 73). According to the results of Structure population analysis, we obtained the optimal population number, with the true number of populations (K) being 3 and the population being divided concerning Q≥0.5. Three subgroups, i.e., Q1, Q2 and Q3, included 34, 33 and 28 germplasms, respectively, and the remaining 10 germplasms were identified as the mixed population. During the experiment, 9 pairs of core primers were screened among the total of 12 for a complete differentiation regarding 105 tested materials, and the fingerprints of 91 C.×morifolium materials and 14 chrysanthemum-related materials were further constructed. Overall, there were significant genetic differences and rich genetic diversity among C.×morifolium materials, which would shed light on the garden application and variety selection fields of C.×morifolium. The fingerprint database of 105 C.×morifolium varieties and chrysanthemum-related species may provide technical support for future research regarding the identification and screening system of C.×morifolium varieties.

Research progress on chemical constituents and pharmacological effects of Ajania plants / 中国中药杂志

Ajania belonging to the subtribe Artemisiinae of Anthemideae(Asteraceae) is a genus of semi-shrubs closely related to Chrysanthemum. There are 24 species of Ajania in northwestern China, most of which are folk herbal medicines with strong stress tolerance. Modern medical studies have demonstrated that the chemical constituents of Ajania mainly include terpenoids, flavonoids, phenylpropanoids, alkynes, and essential oils. These compounds endow the plants with antimicrobial, anti-inflammatory, antitumor, antimalarial, antioxidant, and insecticide effects. In this study, we reviewed the research progress in the chemical constituents and pharmacological activities of Ajania, aiming to provide reference for the further research and development of Ajania.

Correlations between content of linarin in Chrysanthemum indicum and climatic factors in habitats / 中国中药杂志

Chrysanthemi Indici Flos(CIF), the capitulum of Chrysanthemum indicum, is widely used in proprietary Chinese medicine and daily chemical products. At present, CIF is mainly produced from wild resources and rarely cultivated. This study aims to reveal the correlations between linarin content in CIF and climatic factors in different habitats, and provide a theoretical basis for suitable zoning and rational production of medicinal materials. The content of linarin in CIF was determined by HPLC. Grey relational analysis and Pearson correlation analysis were carried out for linarin content with climatic factors. The results showed that the content of linarin in CIF was significantly different among different habitats. The grey relational degrees of climatic factors with linarin content was in an order of average annual precipitation>annual average sunshine hours>annual average temperature>longitude>annual frost-free period>latitude>altitude. Longitude, annual average temperature and average annual precipitation had significantly positive correlations with the content of linarin in CIF, whereas latitude and altitude showed negative correlations with it. The annual frost-free period and annual average sunshine hours had no significant correlation with the content of linarin in CIF. The content of linarin in CIF varied significantly in different habitats. High longitude, low latitude, low altitude, high annual average temperature and high annual average precipitation could be used as indicators for the habitats of high-quality Ch. indicum. This study provides a reference for selecting suitable producing areas of Ch. indicum and establishing artificial cultivation system.

Study on drug-target binding kinetics profiles of flavonoids in Chrysanthemum morifolium and xanthine oxidase / 中国中药杂志

Based on the target occupancy mathematical model, the binding kinetic process of potential active ingredients of lowering uric acid in Chrysanthemum morifolium with xanthine oxidase(XOD) was evaluated. The potential active ingredients of lowering uric acid in Ch. morifolium were screened by UPLC-Q-Exactivems MS technology, reference substance identification and in vitro enzymatic kinetics experiments. The binding kinetic parameters of xanthine oxidase and potential inhibitor in Ch. morifolium were determined by surface plasma resonance(SPR). The verified mathematical model of the XOD target occupancy evaluated the kinetic binding process of inhibitors and xanthine oxidase in vivo. According to UPLC-Q-Exactive MS and reference substance identification, 39 potential uric acid-lowering active ingredients in Ch. morifolium extracts were identified and the inhibitory activities of 23 compounds were determined. Three potential xanthine oxidase inhibitors were screened, namely genistein, luteolin, and apigenin. whose IC_(50 )were 1.23, 1.47 and 1.59 μmol·L~(-1), respectively. And the binding rate constants(K_(on)) were 1.26×10~6, 5.23×10~5 and 6.36×10~5 mol·L~(-1)·s~(-1), respectively. The dissociation rate constants(K_(off)) were 10.93×10~(-2), 1.59×10~(-2), and 5.3×10~(-2 )s~(-1), respectively. After evaluation by different administration methods, the three selected compounds can perform rapid and sustained inhibition of xanthine oxidase in vivo under combined administration. This study comprehensively evaluated the target occupancy process of three effective components in different ways of administration in vivo by UPLC-MS, concentration-response method, SPR technology and xanthine oxidase target occupancy model, which would provide a new research idea and method for screening active ingredients in traditional Chinese medicine.

Research progress of pesticide residues in Chrysanthemum / 中国中药杂志

Chrysanthemum is widely used as a type of edible flower and also considered as the important materials of many beverages in China. Due to the occurrence of diseases and pests, and the lack of regulations for species, frequency, dose of pesticides in Chrysanthemum, pesticides have become one of the main pollutants in Chrysanthemum. The pesticide residues in Chrysanthemum were detected frequently and worth noting. This paper focused on the types of pesticides, pesticide residue detection techniques, and risk assessment methods for Chrysanthemums on the basis of relevant literatures. The pesticide residues of traditional Chinese medicine are mainly organochlorines, organophosphorus and pyrethroids, and the detection techniques include gas chromatography(GC), liquid chromatography(LC) or both combined with mass spectrometry(MS). With the increasing use of traditional Chinese medicine, Chrysanthemum is widely circulated in the market. Therefore, it is important to understand the current situation of pesticide residues in different varieties of Chrysanthemum, so as to provide theoretical reference for the control of quality and safety of Chrysanthemum and the formulation of the maximum residue limit.

Content characteristics of mineral elements and its correlation with soil nutrients and effective components in Chrysanthemum morifolium from different producing areas in Macheng / 中国中药杂志

In this study, 24 copies of samples of Chrysanthemum morifolium and soil from two main production towns in Macheng city were collected, and the contents of 13 mineral elements, 5 effective components and 14 soil nutrient factors in Ch. morifolium were determined. The enrichment characteristics of available soil nutrients by mineral elements were analyzed and the dominant factors affecting the effective components of Ch. morifolium were screened. The results showed that the content of mineral elements and soil nutrients and effective components are very different, and variation of soil fertility was much greater than that of inorganic elements in chrysanthemum plants. In general, the level of element content in Ch. morifolium from different producing areas is K>N>P>Mg>Ca>Fe>Mn>Zn>Cu>Ni>Cr>Pb>Cd. The content of K, N and Mg is higher than that of common crops, and the content of Cu, Cd and Pb in Ch. morifolium from various producing areas does not exceed the relevant standards. The N, P and K enrichment capacity in soil was stronger than that of other elements, and the Ca enrichment ability was the worst. The content of AvCu in the soil was positively correlated with the contents of N, Mg, K, Fe and Cu elements in Ch. morifolium. The contents of chlorogenic acid, luteolin, 3,5-O-dicaffeoylquinic acid reached the pharmacopoeia standard. The percentage of chlorogenic acid and 3,5-O-dicaffeoylquinic acid in Ch. morifolium that from Huangtugang town in the active components were generally higher than that from Futianhe town, and the diffe-rences of luteolin contents in the two producing areas were relatively small. The correlation and regression analysis showed that the contents of Cu, Zn and Cr in Ch. morifolium were positively correlated with the active components, while the contents of Fe, Mn and Ni were negatively correlated with the contents of AvP, AvK, TK, AvMn and AvCu in soil. In general, Zn and Ca fertilizer should be added to the ecological planting of Ch. morifolium, K fertilizer should be added, and N and P fertilizer should be applied appropriately.

Analysis and evaluation of mineral elements of Chrysanthemum morifolium for medicinal and tea use of different germplasm resources / 中国中药杂志

In this study, 23 germplasm resources of Chrysanthemum morifolium used in medicine and tea were collected from Dabie Mountains and its surrounding producing areas, and the contents of 13 mineral elements were determined and compared. The thermal maps of correlation analysis, principal component analysis and cluster analysis were used for comprehensive evaluation. The results showed that the average content of each element in Ch. morifolium of different germplasm resources was: K>N>P>Mg>Ca>Fe>Mn>Zn>Cu>Ni>Cr>Pb>Cd, and the leaves were: K>N>Ca>Mg>P>Fe>Mn>Zn>Cr>Cu>Ni>Pb>Cd. There are rich contents of N, P, K, Ca, Mg and Fe in Ch. morifolium flowers and their leaves, among them, K element has the largest change range, while N, Ca, Fe, Mg and Zn elements have a larger change range. The absorption and accumulation of each element in the leaves of different germplasm resources varied greatly. The correlation analysis shows that there is a strong positive correlation between Ca element, Mg, Mn and Cd element.Principal component analysis in Ch. morifolium flowers characteristic elements for Mn, Cr, Cu, P, K, can be used as a Ch. morifolium resources to identify the characteristics of the elements, choose top five principal component(F1-F5) comprehensive evalua-tion of medicinal Ch. morifolium, scored in the top five varieties for Hangiu-Fuhuangju, Hangju-Xiaoyangju, Hangju-Sheyangju, Hangju-Dayanghua, Hangju-Subeiju,indicates that in terms of mineral elements, the five medicinal Ch. morifolium resources quality is better. The PCA score chart can divide 23 Ch. morifolium resources into 4 groups, and the cluster analysis heat map divides 23 Ch. morifolium resources into 5 groups. All the Ch. morifolium resources of the same type can be well clustered together, indicating that the difference in mineral element content of Ch. morifolium germplasm resources is closely related to genetic factors.

Comprehensive antioxidant and anti-inflammatory activity of alcohol extracts from Chrysanthemum indicum in different areas based on entropy weight and TOPSIS methodology / 中国中药杂志

The study is aiming at investigating the application of entropy weight TOPSIS method in the comparison of the scavenging effect of DPPH, ABTS and hydroxyl radical and the inhibition effect of xanthine oxidase(XOD) and lipoxygenase(LOX) of Chrysanthemum indicum. The DPPH, ABTS, salicylic acid and spectrophotometry were used to determine the scavenging effect of DPPH, ABTS and hydroxyl radical and the inhibition effect of xanthine oxidase(XOD) and lipoxygenase(LOX) of Ch. indicum from 31 different areas in vitro. Take the half inhibition rate of as the evaluation index, two principal components were extracted by the principal component analysis, and their cumulative contribution rate reached at 92.4%. The different areas of Ch. indicum could be divided into Dabei Mountain and Qinling-Taihang Mountain by use principal component to analysis. The entropy weight TOPSIS method was used to objectively assign weights to five indexes, calculate the weight of each index and set up the best and worst scheme of the evaluation object, and the relative proximity(C_i) was used as the measure to construct the multi-index comprehensive evaluation model of Ch. indicum. And then sort with the relative proximity value. The results showed that the relative proximity was between 0.098 and 0.983 which represents there were significant differences in the scavenging effect of DPPH, ABTS and hydroxyl radical and the inhibition effect of xanthine oxidase(XOD) and lipoxygenase(LOX) between extracts of Ch. indicum from different areas. The Ch. indicum from Dabie Mountain area have a relatively high relative degree of measurement and high-quality ranking. Taken together, the quality of Ch. indicum.from the Dabie Mountain area is better. The index weight coefficient and the classification result of producing area are basically consistent with the result of principal component analysis. The results show that the TOPSIS method based on entropy weight method can be used to evaluate the comprehensive quality of Ch. indicum.

Ectopic expression of Chrysanthemum CDM19 in Arabidopsis reveals a novel function in carpel development

BACKGROUND: APETALA3 (AP3) has significant roles in petal and stamen development in accordance with the classical ABC model. RESULTS: The AP3 homolog, CDM19, from Chrysanthemum morifolium cv. Jinba was cloned and sequenced. Sequence and phylogenetic analyses revealed that CDM19 is of DEF/AP3 lineage possessing the characteristic MIKC-type II structure. Expression analysis showed that CDM19 was transcribed in petals and stamens of ray and disc florets with weak expression in the carpels. Ectopic expression of CDM19 in Arabidopsis wild-type background altered carpel development resulting in multi-carpel siliques. CDM19 could only partially rescue the Arabidopsis ap3­­3 mutant. CONCLUSIONS: Our results suggest that CDM19 may partially be involved in petal and stamen development in addition to having novel function in carpel development.

Changes in chemical compositions of Chrysanthemi Flos after frying and protective effects on CCl_4-induced acute liver injury in mice / 中国中药杂志

To reveal the processing mechanism of Chrysanthemi Flos from the changes of chemical compositions after frying and its effect on the efficacy of liver protection. Ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry(UPLC-Q-TOF-MS) and ultra high performance liquid chromatography(HPLC) were used for the qualitative and quantitative researches of chemical compositions before and after Chrysanthemi Flos frying. Progenesis QI and SPSS software were used for principal component analysis(PCA), partial least squares discriminant analysis(PLS-DA), variable importance projection(VIP) analysis and t-test to identify the compositions with significant changes. Pharmacodynamics experiment was used to investigate the protective effect of crude and fried Chrysanthemi Flos on CCl_4-induced acute liver injury in mice. According to mass spectrometry data, there were 28 chemical compositions in crude and fried Chrysanthemi Flos, mainly including flavonoids and organic acids. 13 compositions such as luteolin, apigenin and luteolin glycoside were increased significantly after frying, while 7 compositions such as chlorogenic acid, luteolin-7-O-glucuronide and apigenin-7-O-glucuronide were decreased significantly after frying. Through principal component analysis, crude and fried Chrysanthemi Flos products were divided into two categories, indicating that there were internal differences in quality. The results of liver injury protection experiment in mice showed that the AST, ALT and MDA contents were significantly decreased and SOD level was increased in mice with liver injury in both the high and medium dose groups. Histopathological examination showed that crude and fried Chrysanthemi Flos can protect the liver by reducing inflammatory cell infiltration, reducing steatosis, and repairing damaged liver cells. The results of this study showed that the chemical compositions had obvious changes after frying, and both crude and fried Chrysanthemis Flos had protective effects on CCl_4-induced acute liver injury in mice. In addition, in the range of high, medium and low doses, the liver protection effect of crude and fried Chrysanthemi Flos increased with the increase of dose. The experiment results provided reference for the mechanism of fried Chrysanthemi Flos and clinical selection of processed products.

Effects of planting period on growth, yield and medicinal quality of Chrysanthemum morifolium / 中国中药杂志

In this paper, Chrysanthemum morifolium was used as the experimental object, eight different planting periods were set up in field plot experiment from April to August, which were 04-15, 05-19, 05-30, 06-09, 06-19, 07-20, 07-31, 08-15. The effects of different treatments on the occurrence of root rot, agronnmic traits, mineral element absorption and content of effective components of Ch. morifolium in Macheng country of Hubei province were studied. The results showed that delaying the planting time could effectively reduce the diseases occurrence of root rot of Ch. morifolium. With the advance of transplanting period, the plant height, the weight of one hundred flowers and the number of flowers of Ch. morifolium showed a trend of gradual decrease, while the number of primary branches and the thickness of main stem and the primary branch increased first and then decreased. The yield of Ch. morifolium per plant and per mu increased with the advance of the planting period, and the yield per mu increased during the planting period on June 19, which was 91.96% higher than that on April 15. And with the delay of the planting period,the absorption and accumulation of potassium(K) elements was promotes. The content of active ingredients such as chlorogenic acid, rutin, luteolin, and 3,5-O dicoffeoacy lquinic acid in the Ch. morifolium increased significantly and then gradually decreased with the delay of the planting period, which indicates that late planting can significantly improve the quality of Ch. morifolium. Considering factors such as the occurrence of root rot disease, yield and active ingredient content, combined with climatic conditions in the Dabie Mountains in eastern Hubei, the optimum planting period of Ch. morifolium was determined from mid-late June to early July.

Study on wild tending of Chrysanthemum indicum / 中国中药杂志

To provide theoretical basis for the formation of Chrysanthemum indicum resource and quality,the differences in biology,ecology,yield and quality of Ch. indicum population in the wild tending,the cultivated and the wild were analyzed and compared in this study. The results showed that in the aspect of biology,there was no significant difference between the condition of wild tending and cultivated Ch. indicum at the height of plant,both were significantly higher than wild Ch. indicum. In the aspect of ecology,in the condition of wild tending is less likely to occur water logging than cultivated Ch. indicum. In the condition of wild tending is less likely to occur plant diseases and insect pests than Ch. indicum in the condition of wild. In the aspect of production,the average output of the sample of the condition of wild tending was significantly higher than that the condition of artificial cultivation and wild quadrat. There was no significant difference between the three kinds of condition on the traits of capitulum dichasium,the content of buddleoside of the wild tending Ch. indicum was significantly higher than that of 0. 80% required by the Chinese Pharmacopoeia( 2015 edition). Therefore,the artificial supplement and the appropriate artificial management of wild tending is a mode of production to increase the population density of Ch. indicum and obtain high quality and high yield medicinal materials.

Effect of Chrysanthemum indicum on absorption characteristics of Cd and its effect on quality of medicinal materials / 中国中药杂志

The Cd stress of Chrysanthemum indicum was treated by different concentrations of Cd Cl2 solution in the culture substrate. The content of Cd in different parts of Ch. indicum and the content of buddleoside and the total flavonoids in Ch. indicum were determined. The absorption characteristics of Cd elements in Ch. indicum were analyzed. And the influence of Cd elements on the quality of the herbs. The results showed that the application of soil Cd in the range of 0-100 mg·kg~(-1) had no significant effect on the biomass of Ch. indicum,and the root-shoot ratio showed a decreasing-increasing-decreasing trend. The content of Cd in different parts of Ch. indicum was significantly different,and the content of aboveground part was higher than that of underground part. The enrichment factors of Cd elements in different parts of Ch. indicum are different. The enrichment coefficient of aboveground parts is larger than that of underground parts. The whole parts and plants show an increase first and then decrease,and the overall enrichment factor is greater than1. The transfer coefficient of the aerial part/underground part of Ch. indicum showed a decreasing-increasing-decreasing-increasing trend with the increase of the amount of Cd applied in the soil,and the transfer coefficient was higher than 1. The contents of buddleoside and total flavonoids in Ch. indicum after Cd stress treatment were lower than the control,and the overall performance was lower and then increased,but it was still significantly lower than the control,indicating that Cd pollution directly led to the decrease of chemical quality of Ch. indicum.

Effect of short-day treatment on expression of CO gene in Chrysanthemum indicum / 中国中药杂志

This experiment studied the expression pattern of key gene CO in the photoperiod of Chrysanthemum indicum. The CDS sequence of the Ch. indicum CO gene was cloned by RT-PCR. The open reading frame was 1 380 bp in length and encoded 459 amino acids. The bioinformatics analysis results showed that the Ch. indicum CO had higher homology with Ch. lavandulifolium and Artemisia annua,and the CO was more conservative in the same family. The molecular weight of the predicted protein encode by CO is 52. 04 k Da,the p I is 4. 81,the α-helix structure accounted for 17. 65%,the random coil accounted for 76. 69%,the extension chain accounted for 5. 66%,there are no β-fold and signal peptide. The experimental results showed that short-day treatment could increase the expression level of CO gene in Ch. indicum and induce its flowering. The results of qRT-PCR showed that the relative expression of CO gene in different tissues and different treatment periods of Ch. indicum was significantly different. In this paper,we studied the effect of short-day treatment on the expression of key genes in the flowering cycle of Ch. indicum,providing a basis for photoperiod regulation and harvesting period of Ch.indicum.

Identification of Chrysanthemum indicum and its adulterants based on ITS2 barcode / 中国中药杂志

DNA barcode technology was used to establish a rapid identification method of Chrysanthemum indicum based on ITS2 sequences. The total DNA was extracted from 22 collected samples,and the ITS2 sequence was amplified by PCR and sequenced,and the information of ITS2 sequence was obtained. Another 14 items of the same family or the same genus were downloaded from Gen Bank.We aligned all 36 sequences,calculated the intraspecific and interspecific distances,and constructed Neighbor Joining( NJ) phylogenetic tree,using MEGA 7. 0. The difference of the secondary structure between the ITS2 sequences was compared. The results showed that the genetic distance of Ch. indicum and Ch. morifolium was overlapped,but the maximum intraspecific distance was far less than the minimum interspecific distance between and among Ch. indicum and other species,with an obvious barcoding gap. The NJ tree showed that Ch. indicum and Ch. morifolium shared a clade,and most of Ch. morifolium with some Ch. indicum were shared a subclade,while Inula lineariifolia,Sinosenecio oldhamianus and Senecio scandens belonged to one clade separately. ITS2 secondary structures for I. lineariifolia,S. oldhamianus and S. scandens were significantly different enough to identify completely but Ch. indicum and Ch. morifolium shared two secondary structures of A and B. It was proved that Ch. indicum was one of the evolutionary sources of Ch.morifolium. Therefore ITS2 sequence as DNA barcode can identify Ch. indicum and its adulterants accurately and quickly. The study provides an important basis for Ch. indicum for the identification of germplasm resources and the safety of clinical medication.

Identification of Chrysanthemum indicum in different geographical populations and Ch. morifolium based on DNA barcodes of psbA-trnH,matK and trnL / 中国中药杂志

DNA barcode technology was used to establish a rapid identification method of Chrysanthemum indicum and Ch. morifolium based on psbA-trn H,mat K and trn L sequences. The total DNA was extracted from 21 samples collected,and the psbA-trn H,mat K,trn L sequences were amplified by PCR and sequenced. The information of these sequences were obtained. We aligned all 63 sequences,calculated the intraspecific and interspecific distances,analysed the SNPs distribution of psbA-trn H+mat K+trn L combination sequences and constructed the Neighbor-joining( NJ) Tree,using MEGA 7. 0. The results showed that the genetic distances of Ch. indicum,Ch. indicum( Juhuanao)and Ch. morifolium were overlapped. The SNPs analysis of psbA-trn H+mat K+trn L combination sequences showed that there were 19 nucleotide polymorphism loci( SNPs) and nine parsim-informative sites in the combination sequences. In addition,Ch. indicum showed more obvious sequence polymorphism than those of Ch. indicum( Juhuanao) and Ch. morifolium. The psbA-trn H sequences showed obvious length variation.The NJ Tree showed that Ch. morifolium numbered C2-C5 were clustered into a single subbranch with a bootstrap value of 62%,and Ch.morifolium could be distinguished from Ch. indicum and Ch. indicum( Juhuanao). Moreover,Ch. indicum numbered Z9 and Z10 collected from Gansu province were singly clustered into one branch with a bootstrap value of 77%. It was also found that the changes of psbA-trn H and trn L sequences information of Ch. indicum samples from the northwest were obviously related to the geography and environment. Moreover,Ch.indicum and Ch. indicum( Juhuanao) had obvious differentiation,were also regarded as the evolutionary sources of Ch. morifolium. Therefore,psbA-trn H+mat K+trn L combination sequences as DNA barcode can identify Ch. indicum and Ch. morifolium accurately and rapidly,which provides an important basis for germplasm resources identification and species identification.

Cloning and prokaryotic expression of CHI in Chrysanthemum morifolium cv.'Hangju' / 中国中药杂志

Three Chrysanthemum-chalcone-isomerase genes( CmCHI) were successfully cloned by PCR from the database of Chrysanthemum transcriptome and named CmCHI1,CmCHI2 and CmCHI3,respectively. Bioinformatics analysis showed that the base numbers of CmCHI1-3 open reading frame were 708,633 and 681 bp,encoding 235,210 and 226 amino acids,respectively. Three fusion proteins of about 30 kDa were successfully induced by prokaryotic expression technology,and the corresponding recombinant fusion proteins were isolated and purified by Ni-NTA resin column. Clustering analysis showed that the 3 CmCHI were homologous with Compositae plants,and CmCHI1 and CmCHI3 belonged to type Ⅰ CHI. CmCHI2 belongs to type Ⅳ CHI. Using β-actin as an internal reference gene,RT-qPCR was used to detect and analyze the expression of CmCHI1-3 genes in Hangju. The results showed that the expression levels of CmCHI1 and CmCHI3 were higher,while the expression levels of CmCHI2 were lower. It was concluded that CmCHI1 and CmCHI3 were the main chalcone isomerase genes involved in the synthesis of flavonoids in Hangju,and CmCHI2 was a helper gene. Flooding treatment significantly promoted the expression of CmCHI1 and CmCHI3 genes,but had no regulatory effect on CmCHI2. The above results provided a basis for further study of the molecular regulation mechanism of CHI gene in the metabolism of flavonoids in Hangju,which laid a foundation for improving the content of flavonoids in Hangju and finally improving the medicinal quality of Hangju.

Establishment of HPLC fingerprint and comparison of active ingredients between varieties of chrysanthemum in Hubei province / 中国中药杂志

In this paper,the fingerprint of different varieties of chrysanthemum were established with " Similarity Evaluation System for Chromatographic Fingerprint of Chinese Materia Medica" and the content of chlorogenic acid,galuteolin and 3,5-O-dicaffeoylquinic acid in 29 batches of different varieties of chrysanthemum in Futianhe town,Huangtugang town and Wuhan city were compared. At the same time,similarity evaluation and common peak clustering analysis were carried out. There were 11 common peaks in the fingerprints of 29 batches of different varieties of chrysanthemum,and the similarity ranged from 0. 802 to 0. 975. Hangju and Gongju were divided into one group by cluster analysis,and Huangju into another category. The established fingerprint method provides a basis for the identification of chrysanthemum cultivars. The content of 29 batches of chlorogenic acid was between 4. 092 and 11. 723 mg·g-1,luteolin was between 1. 010 and 11. 713 mg·g-1,and 3,5-O-dicaffeoylquinic acid was between 8. 828 and 33. 435 mg·g-1,both reach the pharmacopoeia standard,but the effective components of different varieties of chrysanthemum were quite different. Based on the contents of three active ingredients and the diversity of fingerprint peaks,the quality of the characteristic germplasm resource of local Fubaijuin Macheng is superior,and the protection of local characteristic germplasm resource should be strengthened in production.

Development of a Molecular Marker for Fruiting Body Pattern in Auricularia auricula-judae

The fruiting body pattern is an important agronomic trait of the edible fungus Auricularia auricula-judae, and an important breeding target. There are two types of fruiting body pattern: the cluster type and the chrysanthemum type. We identified the fruiting body pattern of 26 test strains, and then constructed two different near-isogenic pools. Then, we developed sequence characterized amplified region (SCAR) molecular markers associated with the fruiting body pattern based on sequence-related amplified polymorphism (SRAP) markers. Ten different bands (189–522 bp) were amplified using 153 pairs of SRAP primers. The SCAR marker “SCL-18” consisted of a single 522-bp band amplified from the cluster-type strains, but not the chrysanthemum strains. This SCAR marker was closely associated with the cluster-type fruiting body trait of A. auricula-judae. These results lay the foundation for further research to locate and clone genes controlling the fruiting body pattern of A. auricula-judae.

Morphology and Molecular Characterization of Alternaria argyranthemi on Chrysanthemum coronarium in China

Chrysanthemum coronarium is an economically important plant in Asia, and used medicinally, ornamentally and as a vegetable. In April 2017, leaf spot disease on C. coronarium was observed in Shiyan, Hubei, China. A single-spore isolate was obtained and identified based on morphology and sequence analysis using four regions (rDNA ITS, GAPDH, EF-1α, and RPB2). The results indicated that the fungus is Alternaria argyranthemi. The pathogenicity tests revealed that the species could cause severe leaf spot and blight disease on the host. This is the first report of leaf spot disease on C. coronarium caused by A. argyranthemi in the world, which is also a new record of Alternaria species in China.