RESUMEN

Lonicera macranthoides, the main source of traditional Chinese medicine Lonicerae Flos, is extensively cultivated in Southwest China. However, the quality of L. macranthoides produced in this region significantly varies due to its wide distribution and various cultivation breeds. Herein, 50 Lonicerae Flos samples derived from different breeds of L. macranthoides cultivated in Southwest China were collected for quality evaluation. Six organic acids and three saponin compounds were quantitatively analyzed using HPLC. Furthermore, the antioxidant activity of a portion of samples was conducted with 2,2'-Azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS) and 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenging experiments. According to the quantitative results, all samples met the quality standards outlined in the Chinese Pharmacopoeia. The samples from Guizhou, whether derived from unopened or open wild-type breeds, exhibited high quality, while the wild-type samples showed relatively significant fluctuation in quality. The samples from Chongqing and Hunan demonstrated similar quality, whereas those from Sichuan exhibited relatively lower quality. These samples demonstrated significant abilities in clearing ABTS and DPPH radicals. The relationship between HPLC chromatograms and antioxidant activity, as elucidated by multivariate analysis, indicated that chlorogenic acid, isochlorogenic acid A, isochlorogenic acid B, and isochlorogenic acid C are active components and can serve as Q-markers for quality evaluation.

Asunto(s)

RESUMEN

Due to the medicinal importance of the flowers of Xianglei type (XL) Lonicera macranthoides, it is important to understand the molecular mechanisms that underlie their development. In this study, we elucidated the transcriptomic and metabolomic mechanisms that underlie the flower development mechanism of two L. macranthoides varieties. In this study, 3435 common differentially expressed unigenes (DEGs) and 1138 metabolites were identified. These common DEGs were mainly enriched in plant hormone signal transduction pathways. Metabolomic analysis showed that amino acids were the main metabolites of differential accumulation in wild-type (WT) L. macranthoides, whereas in XL, they were flavonoids and phenylalanine metabolites. Genes and transcription factors (TFs), such as MYB340, histone deacetylase 1 (HDT1), small auxin-up RNA 32 (SAUR32), auxin response factor 6 (ARF6), PIN-LIKES 7 (PILS7), and WRKY6, likely drive metabolite accumulation. Plant hormone signals, especially auxin signals, and various TFs induce downstream flower organ recognition genes, resulting in a differentiation of the two L. macranthoides varieties in terms of their developmental trajectories. In addition, photoperiodic, autonomous, and plant hormone pathways jointly regulated the L. macranthoides corolla opening. SAUR32, Arabidopsis response regulator 9 (ARR9), Gibberellin receptor (GID1B), and Constans-like 10 (COL10) were closely related to the unfolding of the L. macranthoides corolla. These findings offer valuable understanding of the flower growth process of L. macranthoides and the excellent XL phenotypes at the molecular level. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-024-04019-1.

RESUMEN

This study aims to explore the molecular regulatory mechanism of the differential accumulation of flavonoids between 'Xianglei' and the wild type of Lonicera macranthoides. The flowers, stems, and leaves of the two varieties of L. macranthoides were collected. Ultra-performance liquid chromatography-mass spectrometry(UPLC-MS) and high-throughput sequencing(RNA-seq) were employed to screen out the differential flavonoids, key differentially expressed genes(DEGs) and transcription factors(TFs). Fourteen DEGs were randomly selected for verification by qRT-PCR. The results showed that a total of 17 differential flavonoids were obtained, including naringin chalcone, apigenin, and quercetin. The transcriptomic analysis predicted 19 DEGs associated with flavonoids, including 2 genes encoding chitin synthase(CHS) and 3 genes encoding chalcone isomerase(CHI). The regulatory network analysis and weighted gene co-expression network analysis(WGCNA) screen out the key enzyme genes CHS1, FLS1, and HCT regulating the accumulation of flavonoids. MYB12 and LBD4 may be involved in the biosynthesis of flavonoids by regulating the expression of key enzyme genes CHS1, FLS1, and HCT. The qRT-PCR and RNA-seq results were similar regarding the expression patterns of the 14 randomly selected DEGs. This study preliminarily analyzed the transcriptional regulatory mechanism for the differential accumulation of flavonoids in the two varieties of L. macranthoides and laid a foundation for further elucidating the regulatory effects of key enzyme genes and TFs on the accumulation of flavonoids.

Asunto(s)

RESUMEN

BACKGROUND: Flavonoids are one of the bioactive ingredients of Lonicera macranthoides (L. macranthoides), however, their biosynthesis in the flower is still unclear. In this study, combined transcriptomic and targeted metabolomic analyses were performed to clarify the flavonoids biosynthesis during flowering of L. macranthoides. RESULTS: In the three sample groups, GB_vs_WB, GB_vs_WF and GB_vs_GF, there were 25, 22 and 18 differentially expressed genes (DEGs) in flavonoids biosynthetic pathway respectively. A total of 339 flavonoids were detected and quantified at four developmental stages of flower in L. macranthoides. In the three sample groups, 113, 155 and 163 differentially accumulated flavonoids (DAFs) were detected respectively. Among the DAFs, most apigenin derivatives in flavones and most kaempferol derivatives in flavonols were up-regulated. Correlation analysis between DEGs and DAFs showed that the down-regulated expressions of the CHS, DFR, C4H, F3'H, CCoAOMT_32 and the up-regulated expressions of the two HCTs resulted in down-regulated levels of dihydroquercetin, epigallocatechin and up-regulated level of kaempferol-3-O-(6''-O-acetyl)-glucoside, cosmosiin and apigenin-4'-O-glucoside. The down-regulated expressions of F3H and FLS decreased the contents of 7 metabolites, including naringenin chalcone, proanthocyanidin B2, B3, B4, C1, limocitrin-3,7-di-O-glucoside and limocitrin-3-O-sophoroside. CONCLUSION: The findings are helpful for genetic improvement of varieties in L.macranthoides.

Asunto(s)

RESUMEN

Lonicera macranthoides Hand.-Mazz. is a traditional medicinal plant that is cultivated in Hunan, Yunnan, and Guizhou Provinces in China. In June 2020, a new leaf spot disease was observed on this plant in Longhui County, Shaoyang City, Hunan Province, China, where 14,000 hm2 of L. macranthoides had been planted. About 20% of the total cultivated area exhibited symptoms. Brown spots appeared on the leaves during the early stage and gradually expanded into irregular lesions, which became necrotic and dry. The whole plant withered and died in severe cases. To isolate the pathogen, the infected leaves were collected from different fields and washed with flowing sterile water. The small lesions were then cut and surface sterilized with 75% alcohol for 45 s followed by a 3 min treatment in 3% sodium hypochlorite. The lesions were rinsed five times in sterile water, incubated on potato dextrose agar (PDA) plates and cultured for 3-5 d at 28℃. In total, eleven isolates were obtained, and eight of them were Colletotrichum (isolation frequency 73%). Three representative isolates (JYH1, JYH2, and JYH5) were selected for further study. The fungus grew as circular white colonies, which then became grey. The older colonies looked like cotton and had dense aerial hyphae. The conidia were aseptate, transparent, cylindric, and thin walled, which measured 11.54 to 22.64 × 3.55 to 4.75 µm (n=100). Six genetic regions were amplified and sequenced to further confirm the identity of fungus. They included ß-tubulin (TUB2), the internal transcribed spacer (ITS), actin (ACT), chitin synthase (CHS), calmodulin (CAL) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The sequences were submitted to GenBank (ITS: OQ746331; ON954583; OQ746334; TUB2: OQ772278; ON960155; OQ772279; CHS: OQ772280; ON960156; OQ772281; ACT: OQ772282; ON960157; OQ772283; CAL: OQ772284; ON960158; OQ772285; GAPDH: OQ772286; ON960159; OQ772287). The construction of a 6-gene joint phylogenetic tree analysis showed that the three isolates unambiguously clustered with Colletotrichum kahawae subsp. ciggaro strains C022-1 (GenBank: KJ001120.1, KJ001124.1, KJ001109.1, KJ001102.1, KJ001106.1, KJ001113.1) and R019 (GenBank: JN715847.1, KC860023.1, KC859980.1, KC859954.1, KC859972.1, KC859997.1), which was recently reclassified as C. cigarro (Cabral et al. 2020). Three representative isolates were used for the pathogenicity test on the young leaves of the whole plant. A sterile pin was used to prick the leaf epidermis, and 6 × 6 mm mycelial blocks that had been cultured on PDA for 7 d were placed on the leaf wounds. The controls were treated in the same manner except that sterile blocks of PDA were used. There were three replicates per treatment. All the plants used in the experiment were maintained at 28°C in a climate chamber. There was a 12 h photoperiod, and the chamber was kept at 80% relative humidity. Dark brown spots appeared at the sites of inoculation on the plants after 5 days. All the strains that were re-isolated from the lesions shared the same morphological characteristics and had the same type of colonies as the pathogen Colletotrichum ciggaro. Thus, Koch's postulates were fulfilled. C. ciggaro had been shown to cause anthracnose on Olea europaea L. (Weir et al. 2012), Mangifera indica L. (Ismail et al. 2015), Citrus reticulata L. (Perrone et al. 2016) and Areca catechu L. (Zhang et al. 2020). To our knowledge, this is the first report of C. ciggaro causing anthracnose on L. macranthoides in China and worldwide. This research provides a basis for further research to control epidemics of this disease.

RESUMEN

As a large family of transcription factors, the MYB family plays a vital role in regulating flower development. We studied the MYB family members in Lonicera macranthoides for the first time and identified three sequences of 1R-MYB, 47 sequences of R2R3-MYB, two sequences of 3R-MYB, and one sequence of 4R-MYB from the transcriptome data. Further, their physicochemical properties, conserved domains, phylogenetic relationship, protein structure, functional information, and expression were analyzed. The results show that the 53 MYB transcription factors had different conserved motifs, physicochemical properties, structures, and functions in wild type and 'Xianglei' cultivar of L. macranthoides, indicating their conservation and diversity in evolution. The transcript level of LmMYB was significantly different between the wild type and 'Xianglei' cultivar as well as between flowers and leaves, and some genes were specifically expressed. Forty-three out of 53 LmMYB sequences were expressed in both flowers and leaves, and 9 of the LmMYB members showed significantly different transcript levels between the wild type and 'Xianglei' cultivar, which were up-regulated in the wild type. The results provide a theoretical basis for further studying the specific functional mechanism of the MYB family.

Asunto(s)

RESUMEN

Light has important effects on plant metabolism. However, the relationship between the chlorogenic acid (CGA) content and light in plants remains unclear. Here, we investigated the effects of shading treatment on gene expression and CGA content in Lonicera macranthoides Hand.-Mazz. (LM), a widely used medicinal plant. A total of 1891 differentially expressed genes (DEGs) were obtained in flower buds and 819 in leaves in response to light in shading treatment compared to the control sample by RNA-Seq. After shading treatment, the content of CGA in LM leaves decreased significantly by 1.78-fold, the carotenoid content increased, and the soluble sugar and starch contents significantly decreased. WGCNA and the expression of related genes verified by qRT‒PCR revealed that CGA synthesis pathway enzyme genes form a co-expression network with genes for carbohydrate synthesis, photosynthesis, light signalling elements, and transcription factor genes (TFs) that affect the accumulation of CGA. Through a virus-induced gene silencing (VIGS) system and CGA assay in Nicotiana benthamiana (NB), we determined that downregulation of NbHY5 expression decreased the CGA content in NB leaves. In this study, we found that light provides energy and material for the accumulation of CGA in LM, and light affects the expression of CGA accumulation-related genes. Our results show that different light intensities have multiple effects on leaves and flower buds in LM and are able to coregulate LmHY5 expression and CGA synthesis.

Asunto(s)

RESUMEN

BACKGROUND: Lonicera macranthoides Hand.-Mazz. is an important medicinal plant. Xianglei-type (XL) L. macranthoides was formed after many years of cultivation by researchers on the basis of the natural mutant. The corolla of L. macranthoides XL remains unexpanded and its flowering period is nearly three times longer than that of wild-type (WT) plants. However, the molecular mechanism behind this desirable trait remains a mystery. OBJECTIVE: To understand the floral phenotype differences between L. macranthoides and L. macranthoides XL at the molecular level. METHODS: Transcriptome analysis was performed on L. macranthoides XL and WT. One DEG was cloned by RT-PCR amplification and selected for qRT-PCR analysis. RESULTS: Transcriptome analysis showed that there were 5603 differentially expressed genes (DEGs) in XL vs. WT. Enrichment analysis of DEGs showed that pathways related to plant hormone signal transduction were significantly enriched. We identified 23 key genes in ethylene biosynthesis and signal transduction pathways. The most abundant were the ethylene biosynthesis DEGs. In addition, the open reading frames (ORFs) of WT and XL ETR2 were successfully cloned and named LM-ETR2 (GenBank: MW334978) and LM-XL-ETR2 (GenBank: MW334978), respectively. qRT-PCR at different flowering stages suggesting that ETR2 acts in the whole stage of flower development of WT and XL. CONCLUSIONS: This study provides new insight into the molecular mechanism that regulates the development of special traits in the flowers of L. macranthoides XL. The plant hormone ethylene plays an important role in flower development and flowering duration prolongation in L. macranthoides. The ethylene synthesis gene could be more responsible for the flower phenotype of XL. The genes identified here can be used for breeding and improvement of other flowering plants after functional verification.

Asunto(s)

RESUMEN

As a large family of transcription factors, the MYB family plays a vital role in regulating flower development. We studied the MYB family members in Lonicera macranthoides for the first time and identified three sequences of 1R-MYB, 47 sequences of R2R3-MYB, two sequences of 3R-MYB, and one sequence of 4R-MYB from the transcriptome data. Further, their physicochemical properties, conserved domains, phylogenetic relationship, protein structure, functional information, and expression were analyzed. The results show that the 53 MYB transcription factors had different conserved motifs, physicochemical properties, structures, and functions in wild type and 'Xianglei' cultivar of L. macranthoides, indicating their conservation and diversity in evolution. The transcript level of LmMYB was significantly different between the wild type and 'Xianglei' cultivar as well as between flowers and leaves, and some genes were specifically expressed. Forty-three out of 53 LmMYB sequences were expressed in both flowers and leaves, and 9 of the LmMYB members showed significantly different transcript levels between the wild type and 'Xianglei' cultivar, which were up-regulated in the wild type. The results provide a theoretical basis for further studying the specific functional mechanism of the MYB family.

Asunto(s)

RESUMEN

In order to explore the functions of genes of key rate-limiting enzymes chalcone isomerase(CHI) and chalcone synthase(CHS) in the biosynthesis of flavonoids in Lonicera macranthoides, this study screened and cloned the cDNA sequences of CHI and CHS genes from the transcriptome data of conventional variety and 'Xianglei' of L. macranthoides. Online bioinformatics analysis software was used to analyze the characteristics of the encoded proteins, and quantitative reverse-transcription polymerase chain reaction(qRT-PCR) to detect the expression of CHI and CHS in different parts of the varieties at different flowering stages. The content of luteo-loside was determined by high performance liquid chromatography(HPLC) and the correlation with the expression of the two genes was analyzed. The results showed that the CHI and CHS of the two varieties contained a 627 bp and 1170 bp open reading frame(ORF), respectively, and the CHI protein and CHS protein were stable, hydrophilic, and non-secretory. qRT-PCR results demonstrated that CHI and CHS of the two varieties were differentially expressed in stems and leaves at different flowering stages, particularly the key stages. Based on HPLC data, luteoloside content was in negative correlation with the relative expression of the genes. Thus, CHI and CHS might regulate the accumulation of flavonoids in L. macranthoides, and the specific functions should be further studied. This study cloned CHI and CHS in L. macranthoides and analyzed their expression for the first time, which laid a basis for investigating the molecular mechanism of the differences in flavonoids such as luteoloside in L. macranthoides and variety breeding.

Asunto(s)

RESUMEN

In order to explore the functions of genes of key rate-limiting enzymes chalcone isomerase(CHI) and chalcone synthase(CHS) in the biosynthesis of flavonoids in Lonicera macranthoides, this study screened and cloned the cDNA sequences of CHI and CHS genes from the transcriptome data of conventional variety and 'Xianglei' of L. macranthoides. Online bioinformatics analysis software was used to analyze the characteristics of the encoded proteins, and quantitative reverse-transcription polymerase chain reaction(qRT-PCR) to detect the expression of CHI and CHS in different parts of the varieties at different flowering stages. The content of luteo-loside was determined by high performance liquid chromatography(HPLC) and the correlation with the expression of the two genes was analyzed. The results showed that the CHI and CHS of the two varieties contained a 627 bp and 1170 bp open reading frame(ORF), respectively, and the CHI protein and CHS protein were stable, hydrophilic, and non-secretory. qRT-PCR results demonstrated that CHI and CHS of the two varieties were differentially expressed in stems and leaves at different flowering stages, particularly the key stages. Based on HPLC data, luteoloside content was in negative correlation with the relative expression of the genes. Thus, CHI and CHS might regulate the accumulation of flavonoids in L. macranthoides, and the specific functions should be further studied. This study cloned CHI and CHS in L. macranthoides and analyzed their expression for the first time, which laid a basis for investigating the molecular mechanism of the differences in flavonoids such as luteoloside in L. macranthoides and variety breeding.

Asunto(s)

RESUMEN

BACKGROUND: Lonicera macranthoides is an important woody plant with high medicinal values widely cultivated in southern China. WRKY, one of the largest transcription factor families, participates in plant development, senescence, and stress responses. However, a comprehensive study of the WRKY family in L. macranthoides hasn't been reported previously. OBJECTIVE: To establish an extensive overview of the WRKY family in L. macranthoides and identify senescence-responsive members of LmWRKYs. METHODS: RNA-Seq and phylogenetic analysis were employed to identify the LmWRKYs and their evolutionary relationships. Quantitative real-time (qRT-PCR) and transgenic technology was utilized to investigate the roles of LmWRKYs in response to developmental-, cold-, and ethylene-induced senescence. RESULTS: A total of 61 LmWRKY genes with a highly conserved motif WRKYGQK were identified. Phylogenetic analysis of LmWRKYs together with their orthologs from Arabidopsis classified them into three groups, with the number of 15, 39, and 7, respectively. 17 LmWRKYs were identified to be differentially expressed between young and aging leaves by RNA-Seq. Further qRT-PCR analysis showed 15 and 5 LmWRKY genes were significantly induced responding to tissue senescence in leaves and stems, respectively. What's more, five LmWRKYs, including LmWRKY4, LmWRKY5, LmWRKY6, LmWRKY11, and LmWRKY16 were dramatically upregulated under cold and ethylene treatment in both leaves and stems, indicating their involvements commonly in developmental- and stress-induced senescence. In addition, function analysis revealed LmWRKY16, a homolog of AtWRKY75, can accelerate plant senescence, as evidenced by leaf yellowing during reproductive growth in LmWRKY16-overexpressing tobaccos. CONCLUSION: The results lay the foundation for molecular characterization of LmWRKYs in plant senescence.

Asunto(s)

RESUMEN

In order to study the regulation mechanism of secondary metabolites biosynthesis in Lonicera macranthoides, the key genes involved in the regulation of biosynthesis and the mechanism of differential metabolites were explored. In this study, high-throughput sequencing technology was used for transcriptome sequencing of L. macranthoides at different development stages. By using Liquid chromatography-tandem mass spectrometry(LC-MS/MS) technology, the laws of qualitative, quantitative and synthetic accumulation of its metabolites were studied, and the key enzyme genes for the biosynthesis of phenolic acid and flavonoids were screened out according to the differentially expressed genes. A total of 111 differentially accumulate metabolites(DAM) and 6 653 differentially expressed genes(DGE) were obtained by metabonomics and transcriptomics analysis. The metabolites and key enzyme genes in the Erqing(KE) were significantly different from those in the Dabai(KD) and Yinhua(KY) stages. In the phenylalanine biosynthesis pathway, the ion abundance of chlorogenic acid, naringin, quercetin, rutin, coniferol and other metabolites decreased with the development of flowers, while the ion abundance of ferulic acid, coumarin and syringoside increased with the development of flowers. Key enzyme genes such as CHS, HCT, CCR, FLS and COMT positively regulate the downstream metabolites, while PAL, C4H and 4CL negatively regulate the downstream metabolites. This study provides candidate genes and theoretical basis for the further exploration of key enzymes in the biosynthesis of secondary metabolites and for the regulation of the accumulation of secondary metabolites in L. macranthoides by molecular biological methods.

Asunto(s)

RESUMEN

Lonicera macranthoides Hand-Mazz is an important medicinal plant widely distributed in southern China that has long been used in Chinese traditional medicines. Chlorogenic acid (CGA, 3-caffeoylquinic acid) is the major biologically active ingredient in L. macranthoides. Although key CGA biosynthetic genes have been well documented, their transcriptional regulation remains largely unknown. In this study, we observed that a R2R3 MYB transcription factor LmMYB15 showed a significant correlation with CGA content, indicating its potential role in CGA biosynthesis. A yeast two-hybrid assay suggested that LmMYB15 functions as a transcriptional activator. Overexpression of LmMYB15 in tobacco led to increased accumulation of CGA compared to those in wild-type leaves. To elucidate its functional mechanism, genome-wide DAP-seq was employed and identified the conserved binding motifs of LmMYB15, that is [(C/T) (C/T) (C/T) ACCTA(C/A) (C/T) (A/T)], as well as its direct downstream target genes, including 4CL, MYB3, MYB4, KNAT6/7, IAA26, and ETR2. Subsequently, yeast one-hybrid and dual-luciferase reporter assays verified that LmMYB15 could bind and activate the promoters of 4CL, MYB3 and MYB4, thereby facilitating CGA biosynthesis and phenylpropanoid metabolism. Our findings provide a new track for breeding strategies aiming to enhance CGA content in L. macranthoides that can significantly contribute to better mechanical properties.

Asunto(s)

RESUMEN

A new nitrogen-containing iridoid glycoside, named (7 R,3'R)-lonijapospiroside A (1), together with thirteen known iridoid glycosides, were isolated from the flower buds of Lonicera macranthoides. The structures of these compounds were established on the basis of spectroscopic analyses. Among them, compounds 1-4 are four diastereoisomers, and their absolute configurations were accurately established by the NOE spectra as well as comparison of their experimental and calculated ECD spectra. The anti-inflammatory activities of all isolates were evaluated by measuring their inhibitory effects on NO, IL-6, and TNF-α production in LPS stimulated RAW 264.7 macrophages. Compound 14 exhibited anti-inflammatory activities by inhibiting IL-6 with an IC50 value of 54.70 µM, comparable to that of the positive control (hydrocortisone, IC50: 62.6 ± 1.7 µM).

Asunto(s)

RESUMEN

In order to study the regulation mechanism of secondary metabolites biosynthesis in Lonicera macranthoides, the key genes involved in the regulation of biosynthesis and the mechanism of differential metabolites were explored. In this study, high-throughput sequencing technology was used for transcriptome sequencing of L. macranthoides at different development stages. By using Liquid chromatography-tandem mass spectrometry(LC-MS/MS) technology, the laws of qualitative, quantitative and synthetic accumulation of its metabolites were studied, and the key enzyme genes for the biosynthesis of phenolic acid and flavonoids were screened out according to the differentially expressed genes. A total of 111 differentially accumulate metabolites(DAM) and 6 653 differentially expressed genes(DGE) were obtained by metabonomics and transcriptomics analysis. The metabolites and key enzyme genes in the Erqing(KE) were significantly different from those in the Dabai(KD) and Yinhua(KY) stages. In the phenylalanine biosynthesis pathway, the ion abundance of chlorogenic acid, naringin, quercetin, rutin, coniferol and other metabolites decreased with the development of flowers, while the ion abundance of ferulic acid, coumarin and syringoside increased with the development of flowers. Key enzyme genes such as CHS, HCT, CCR, FLS and COMT positively regulate the downstream metabolites, while PAL, C4H and 4CL negatively regulate the downstream metabolites. This study provides candidate genes and theoretical basis for the further exploration of key enzymes in the biosynthesis of secondary metabolites and for the regulation of the accumulation of secondary metabolites in L. macranthoides by molecular biological methods.

Asunto(s)

RESUMEN

Objective::To clone p-coumaroyl quinate/shikimate 3' -hydroxylase gene from Lonicera macranthoides, and analyze its bioinformatics and expression patterns with chlorogenic acid content, in order to speculate the functions of LmC3H1 gene from L. macranthoides. Method::The full-length cDNA sequence of LmC3H1 gene was cloned by reverse trascription polymerase chain reaction(RT-PCR) and RACE techniques. The bioinformatics analysis of the gene sequence was carried out by using relevant software.Real-time fluorescence quantification PCR(Real-time PCR) and HPLC were used to determine relative expression of LmC3H1 and content of chlorogenic acid in stems, leaves and flowers of different flowering stages. Result::The LmC3H1 (GenBank: MN177695) gene was cloned, and the open reading frame (ORF) of it was 1 533 bp in length and encoded 510 amino acids. The molecular formula was C2618H4134N718O727S22, the relative molecular mass was 58 005.32, and the isoelectric point was 8.92.It was a hydrophilic protein located in the chloroplast with a transmembrane region LLLIPAVLFLISLVYPLI, and contained a conserved domain CYTOCHROME_P450(433-422 aa) in cytochrome P450.The results of Real-time PCR showed that LmC3H1 was expressed in different degrees in stems, leaves and different flowering stages of L. macranthoides. In the flower development stage, the relative expression of white bud stage was the highest, followed by flower buds and white flowering stage. The ratio of flower to stem and leaf was the highest, and the relative expression of flower was the highest. The HPLC results showed that the content of chlorogenic acid increased from greenish white to golden yellow in flowering stage and golden yellow flowering stage. Among the different organs, the flower had the highest chlorogenic acid, and the stem showed the lowest. Conclusion::The LmC3H1 gene of L. macranthoides is cloned, suggesting that LmC3H1 might be involved in the biosynthesis of L. macranthoides chlorogenic acid. This study provides a basis for further studying the functions of the gene and exploring the biosynthesis and regulation mechanism of L. macranthoides chlorogenic acid, while laying the foundation for the genetic improvement of L. macranthoides.

RESUMEN

Objective: To study the glycosides from the 70% ethanol extract of Lonicera macranthoides. Methods: The compounds were isolated and purified by column chromatography of HP-20 macroporous resin, silica gel, ODS, Sephadex LH-20, and semi-preparative RP-HPLC. Their structures were elucidated by physicochemical properties and spectral analyses. Results: Eight compounds were isolated and identified as 7,3’,4’-trimethoxylquercetin-3-O-α-L-arabinadosyl-(1→6)-O-β-D-glucopyranoside (1), 7,3’,4’-trimethoxylquercetin-3-O-rutoside (2), quercetin-3-O-β-D-glucopyranoside (3), (2E,6S)-8-[α-L-arabinopyranosyl-(1″→6’)- β-D-glucopyranosyl]-2,6-dimethyloct-2-eno-1,2″-lactone (4), kankanoside E (5), betulalbuside A (6), shomaside F (7), and amarantholidoside V (8), respectively. Conclusion: Compound 1 is a new compound named methoxylquercetinside, while compounds 5-8 are isolated from the genus of Lonicera for the first time.

RESUMEN

Three previously undescribed compounds, two prenyleudesmanes (1 and 2), and one hexanorlanostane (3), were isolated from the roots of Lonicera macranthoides. Their structures were established based on 1D and 2D nuclear magnetic resonance (NMR) spectra and high-resolution electrospray ionization mass spectral (HR-ESI-MS) data. The absolute configurations of 1 and 3 were determined by X-ray diffraction. To the best of our knowledge, this is the first time that the absolute configuration of a prenyleudesmane with a trans-decalin system and a hexanorlanostane have been unambiguously confirmed by single-crystal X-ray diffraction with Cu Kα radiation. Thecompounds were tested for their antiproliferative activity on the cancer cell lines (HepG2 and HeLa). The compounds 1-3 exhibited moderate inhibitory effects against two human cancer cell lines.

Asunto(s)

RESUMEN

Lonicera macranthoides Hand. -Mazz. is an important medicinal and economical plant in China, however, the codon usage bias (CUB) in L. macranthoides genes is still unknown. In this study, L. macranthoides transcriptome sequencing has been completed, and codon usage patterns in 36,090 reconstructed genes from the L. macranthoides transcriptome were examined. The mean GC content and GC3 value is 44.9% and 43.1%, respectively, which indicates that nucleotide contents of L. macranthoides genome is somewhat AT rich, and its codon bias pattern tends to use A/T-ending codons. According to neutrality plot, ENC plot, PR2-Bias plot and correspondence analysis, we know that both compositional constraint under selection and mutation could affect the CUB in L. macranthoides, and the mutation is the most determinant factor. Meanwhile, gene expression levels can influence its codon usage pattern. Furthermore, we identified 29 optimal codons and most of them ended with A/U. The study will lay a foundation for future research on gene prediction, genetic engineering and molecular evolution in L. macranthoides.

Asunto(s)