Study on chemical constituents and antioxidant activities of Dianthus caryophyllus L.

Objective: Carnation is a plant that holds high value in terms of its edibility, medicinal properties, and ornamental appeal. Creating no sense he aim of this study was to evaluate the antioxidant and antitumor properties of extracts derived from various parts of the carnation plant. Metabolomics technology was employed to identify the primary chemical constituents. Methods: Initially, we measured the total phenolic and total flavonoid contents in carnation roots, stems, leaves, and flowers, followed by assessing the antioxidant and anti-tumor capabilities of each component using diverse experimental methods. Subsequently, UPLC-MS/MS was employed to identify metabolites in different parts of carnation and investigate their roles in antioxidant and anti-tumor activities. Results: Mention numerical value- for better underatnding- Results of the study indicated that the methanol extract obtained from carnation flowers and roots exhibited superior antioxidant capacity compared to that from the stems and leaves. This disparity may be attributed to the abundance of polyphenols, flavonoids, and antioxidants present in the flowers, including methyl ferulate and luteolin-4'-O-glucoside. Furthermore, the significant presence of the anthraquinone compound rhein-8-O-glucoside in carnation roots may contribute to their enhanced antioxidant properties. Ten distinct compounds were isolated and recognized in carnation flowers, with Isoorientin 2"-O-rhamnoside and Kurarinone demonstrating notable antioxidant activity and binding affinity to SOD1 and SOD3, as validated through antioxidant screening and molecular docking. Conclusion: Overall, the findings from this study have expanded our knowledge of the phytochemical composition across different anatomical regions of the carnation plant, providing valuable insights for its holistic utilization.

Novel electroporation-based genome editing of carnation plant tissues using RNPs targeting the anthocyanidin synthase gene.

MAIN CONCLUSION: A novel electroporation method for genome editing was performed using plant tissue samples by direct RNPs-introduction in carnation. Genome editing is becoming a very useful tool in plant breeding. In this study, a novel electroporation method was performed for genome editing using plant tissue samples. The objective was to create a flower color mutant using the pink-flowered carnation 'Kane Ainou 1-go'. For this purpose, a ribonucleoprotein consisting of guide RNA and clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) was introduced into the stem tissue to induce mutations in the anthocyanidin synthase (ANS) gene, which is involved in anthocyanin biosynthesis. As the ANS of 'Kane Ainou 1-go' has not been previously isolated, we initially isolated the ANS gene from 'Kane Ainou 1-go' for characterization. Southern hybridization analysis confirmed that the ANS gene was present in the genome as a two-allele gene with a pair of homologous sequences (ANS-1 and 2); these sequences were used as the target for genome editing. Genome editing was performed by introducing #2_single-guide RNA into the stem tissue using the ribonucleoprotein. This molecule was used because it exhibited the highest efficiency in an analysis of cleavage activity against the target sequence in vitro. Cleaved amplified polymorphic sequence analysis of genomic DNA extracted from 85 regenerated individuals after genome editing was performed. The results indicated that mutations in the ANS gene may have been introduced into two lines. Cloning of the ANS gene in these two lines confirmed the introduction of a single nucleotide substitution mutation for ANS-1 in both lines, and a single amino acid substitution in one line. We discussed the possibility of color change by the amino acid substitution, and also the future applications of this technology.

DNA methylation remodeled amino acids biosynthesis regulates flower senescence in carnation (Dianthus caryophyllus).

Dynamic DNA methylation regulatory networks are involved in many biological processes. However, how DNA methylation patterns change during flower senescence and their relevance with gene expression and related molecular mechanism remain largely unknown. Here, we used whole genome bisulfite sequencing to reveal a significant increase of DNA methylation in the promoter region of genes during natural and ethylene-induced flower senescence in carnation (Dianthus caryophyllus L.), which was correlated with decreased expression of DNA demethylase gene DcROS1. Silencing of DcROS1 accelerated while overexpression of DcROS1 delayed carnation flower senescence. Moreover, among the hypermethylated differentially expressed genes during flower senescence, we identified two amino acid biosynthesis genes, DcCARA and DcDHAD, with increased DNA methylation and reduced expression in DcROS1 silenced petals, and decreased DNA methylation and increased expression in DcROS1 overexpression petals, accompanied by decreased or increased amino acids content. Silencing of DcCARA and DcDHAD accelerates carnation flower senescence. We further showed that adding corresponding amino acids could largely rescue the senescence phenotype of DcROS1, DcCARA and DcDHAD silenced plants. Our study not only demonstrates an essential role of DcROS1-mediated remodeling of DNA methylation in flower senescence but also unravels a novel epigenetic regulatory mechanism underlying DNA methylation and amino acid biosynthesis during flower senescence.

An insertion of transposon in DcNAP inverted its function in the ethylene pathway to delay petal senescence in carnation (Dianthus caryophyllus L.).

Petal senescence is the final stage of flower development. Transcriptional regulation plays key roles in this process. However, whether and how post-transcriptional regulation involved is still largely unknown. Here, we identified an ethylene-induced NAC family transcription factor DcNAP in carnation (Dianthus caryophyllus L.). One allele, DcNAP-dTdic1, has an insertion of a dTdic1 transposon in its second exon. The dTdic1 transposon disrupts the structure of DcNAP and causes alternative splicing, which transcribes multiple domain-deleted variants (DcNAP2 and others). Conversely, the wild type allele DcNAP transcribes DcNAP1 encoding an intact NAC domain. Silencing DcNAP1 delays and overexpressing DcNAP1 accelerates petal senescence in carnation, while silencing and overexpressing DcNAP2 have the opposite effects, respectively. Further, DcNAP2 could interact with DcNAP1 and interfere the binding and activation activity of DcNAP1 to the promoters of its downstream target ethylene biosynthesis genes DcACS1 and DcACO1. Lastly, ethylene signalling core transcriptional factor DcEIL3-1 can activate the expression of DcNAP1 and DcNAP2 in the same way by binding their promoters. In summary, we discovered a novel mechanism by which DcNAP regulates carnation petal senescence at the post-transcriptional level. It may also provide a useful strategy to manipulate the NAC domains of NAC transcription factors for crop genetic improvement.

Low-Oxygen Responses of Cut Carnation Flowers Associated with Modified Atmosphere Packaging.

Gaseous factors affect post-harvest physiological processes in horticultural crops, including ornamental flowers. However, the molecular responses of cut flowers to the low-oxygen conditions associated with modified atmosphere packaging (MAP) have not yet been elucidated. Here, we show that storage of cut carnation flowers in a sealed polypropylene bag decreased the oxygen concentration in the bag to 3-5% and slowed flower opening. The vase life of carnation flowers after storage for seven days under MAP conditions was comparable to that without storage and was improved by the application of a commercial-quality preservative. The adenylate energy charge (AEC) was maintained at high levels in petals from florets stored under MAP conditions. This was accompanied by the upregulation of four hypoxia-related genes, among which the HYPOXIA-RESPONSIVE ETHYLENE RESPONSE FACTOR and PHYTOGLOBIN genes (DcERF19 and DcPGB1) were newly identified. These results suggest that hypoxia-responsive genes contribute to the maintenance of the energy status in carnation flowers stored under MAP conditions, making this gas-controlling technique potentially effective for maintaining cut flower quality without cooling.

Widely targeted metabolomics reveals the antioxidant and anticancer activities of different colors of Dianthus caryophyllus.

Carnation is edible flower that has potent antioxidant properties and is used in traditional Chinese medicinal system and food industry. The phytochemicals responsible for these various proprieties, however, are not fully understood. Thus, in order to recognize metabolite diversity and variability in carnation flowers of different colors and to discover key metabolites that contribute to the differences in antioxidant and anticancer activities, widely targeted LC-MS/MS-based metabolomics analysis was conducted on purple, green, yellow, and white carnation flowers. We identified and chemically categorized 932 metabolites. Metabolic compounds varied significantly with flower color. Several flavonoids, organic acids, phenolic acids, and nucleotides and their derivatives were found to be specific differential metabolites in purple flowers. A total of 128 key differential metabolites were screened. The purple flowers were found to have the highest antioxidant and anticancer activities compared to the other colored flowers. Correlation analysis revealed that the 6-hydroxykaempferol-3,6-O-diglucoside, 6-hydroxykaempferol-7-O-glucoside, quercetin-3-O-sophoroside, and 2'-deoxyguanosine were found to be the major constituents of the antioxidant and anticancer activities. 2'-Deoxyguanosine has effective antiproliferative activity against A549 and U2OS cells for the first report. At the same time, the combination of 2'-deoxyguanosine with 6-hydroxykaempferol-3, 6-O-diglucoside, or quercetin-3-O-sophoroside have also been found to increase the antitumor activity of 2'-deoxyguanosine. These discoveries enrich information on the phytochemical composition of carnation of different colors and provide resources for the overall use and improvement of carnation flowers quality.

Genome-Wide Identification of the MYB and bHLH Families in Carnations and Expression Analysis at Different Floral Development Stages.

Carnations are one of the most popular ornamental flowers in the world with varied flower colors that have long attracted breeders and consumers alike. The differences in carnation flower color are mainly the result of the accumulation of flavonoid compounds in the petals. Anthocyanins are a type of flavonoid compound that produce richer colors. The expression of anthocyanin biosynthetic genes is mainly regulated by MYB and bHLH transcription factors. However, these TFs have not been comprehensively reported in popular carnation cultivars. Herein, 106 MYB and 125 bHLH genes were identified in the carnation genome. Gene structure and protein motif analyses show that members of the same subgroup have similar exon/intron and motif organization. Phylogenetic analysis combining the MYB and bHLH TFs from Arabidopsis thaliana separates the carnation DcaMYBs and DcabHLHs into 20 subgroups each. Gene expression (RNAseq) and phylogenetic analysis shows that DcaMYB13 in subgroup S4 and DcabHLH125 in subgroup IIIf have similar expression patterns to those of DFR, ANS, and GT/AT, which regulate anthocyanin accumulation, in the coloring of carnations, and in red-flowered and white-flowered carnations, DcaMYB13 and DcabHLH125 are likely the key genes responsible for the formation of red petals in carnations. These results lay a foundation for the study of MYB and bHLH TFs in carnations and provide valuable information for the functional verification of these genes in studies of tissue-specific regulation of anthocyanin biosynthesis.

Preparation of carnation-like Ag-ZnO composites for enhanced photocatalysis under visible light.

Carnation-like ZnO was synthesized by the facile precipitation method (at room temperature and in 120 min) to decompose dyes in an aqueous medium. The carnation-like ZnO had a stratified porous structure with a size of about 2-3µm, its petals had a smooth surface with a thickness of 5-10 nm and a width of about 300-500 nm. Ag-ZnO composites were synthesized using glucose with the assistance of PVP. The morphology of Ag-ZnO composites was almost unchanged compared to ZnO. Where, the Ag nanoparticles in the size range of 5-15 nm were uniformly dispersed on the ZnO petals, improving the catalytic ability of the composites in tartrazine (TA) degradation. The influence of Ag content on catalytic structure and performance of composite was studied. The 5Ag-ZnO sample had the highest BET surface area and pore volume and the lowest gap energy (Eg) among the as-synthesized samples. The 5Ag-ZnO sample proclaimed the degradation efficiency in 70 min of 97.8% and thekapof 0.031 min-1. The influences of catalyst content, solution pH, and concentration of dye on the photodegradation efficiency of the composite were thoroughly studied. Besides, the photocatalytic activity of the composite was demonstrated by degrading various organic substances and reusability. In addition, it was compared to a metal-semiconductor catalyst of Au-ZnO and semiconductor-semiconductor catalysts of MoS2-ZnO, Cu2O-ZnO, and SiO2-ZnO. The catalytic mechanism under visible light was proposed.

Exogenous melatonin ameliorates heat damages by regulating growth, photosynthetic efficiency and leaf ultrastructure of carnation.

Carnation (Dianthus caryophyllus L.) is a floral crop that is highly valuable commercially. However, high temperatures adversely affect its growth and the quality of its cut flowers. Melatonin (MT) is a indole substance that can mitigate plant damage under heat stress. In this study, the leaves of carnation seedlings were sprayed with different concentrations of MT before exposure to high temperature. The indices of growth, physiological and chlorophyll fluorescence were measured and analyzed by the membership function method. The results showed that treatment with 100 µM MT was the most effective at ameliorating damage on carnation. We then analyzed the effects of 100 µM MT pretreatment on carnation at different time points of heat stress and found that this concentration of MT ameliorated the damage caused by heat stress, increased the content of photosynthetic pigments, enhanced the performance of photosystem II and improved photosynthesis. In addition, MT also reduced cell damage and lipid peroxidation, increased the activities of antioxidant enzymes and regulated the accumulation of osmotic substances in carnation. Moreover, MT increased the fresh/dry weight of stems and roots, promoted the opening of stomata, and protected the integrity of chloroplast structure of carnation. Compared with heat stress, pre-spraying with MT significantly down-regulated the transcription of a chlorophyll degradation gene and up-regulated the transcription of stress-related genes. Overall, this study provides a theoretical foundation for the mitigation of the adverse effects of exogenous MT under heat stress and proposes beneficial implications for the management of other plants subjected to global warming.

A novel perspective to investigate how nanoselenium and melatonin lengthen the cut carnation vase shelf.

Nano-selenium (nano-Se) and melatonin (MT) applications confirmed to boost plant growth and resistance. The mechanism of various ratios of nano-Se and MT foliar application postpone the senescence of fresh cut carnation flowers and improve vase life remains unclear. In this study, a combined effect with nano-Se (nano-Se5, 5 mg/L) and MT(MT1, 1 mg/L) was preferable to the control, nano-Se, and MT treatment alone when it came to delaying flower senescence. They enhance the antioxidant ability of carnation flowers by lowering MDA and H2O2 levels, raising SOD and POD concentrations, and lowering procyanidins biosynthesis (catechins and epicatechin). Inducing the biosynthesis of hormonal compounds (salicylic acid, jasmonic acid, and abscisic acid), their combination also boosted the growth of carnations. Biofortification with nano-Se and MT substantially increased the amounts of key lignin biosynthesis pathway metabolites (L-phenylalanine, p-hydroxycinnamic acid, p-coumaric acid, perillyl alcohol, p-Coumaryl alcohol, and cinnamic acid), which may increase stem cellular thickness and facilitate water absorption and transmission. The study hypothesizes that nano-Se and MT synergistic applications act as a new efficient non-toxic preservative to extend the vase life and improve the decorative value of carnations.

The mutual regulation between DcEBF1/2 and DcEIL3-1 is involved in ethylene induced petal senescence in carnation (Dianthus caryophyllus L.).

Carnation (Dianthus caryophyllus L.) is a respiratory climacteric flower, comprising one of the most important cut flowers that is extremely sensitive to plant hormone ethylene. Ethylene signaling core transcription factor DcEIL3-1 plays a key role in ethylene induced petal senescence in carnation. However, how the dose of DcEIL3-1 is regulated in the carnation petal senescence process is still not clear. Here, we screened out two EBF (EIN3 Binding F-box) genes, DcEBF1 and DcEBF2, which showed quick elevation by ethylene treatment according to the ethylene induced carnation petal senescence transcriptome. Silencing of DcEBF1 and DcEBF2 accelerated, whereas overexpression of DcEBF1 and DcEBF2 delayed, ethylene induced petal senescence in carnation by influencing DcEIL3-1 downstream target genes but not DcEIL3-1 itself. Furthermore, DcEBF1 and DcEBF2 interact with DcEIL3-1 to degrade DcEIL3-1 via an ubiquitination pathway in vitro and in vivo. Finally, DcEIL3-1 binds to the promoter regions of DcEBF1 and DcEBF2 to activate their expression. In conclusion, the present study reveals the mutual regulation between DcEBF1/2 and DcEIL3-1 during ethylene induced petal senescence in carnation, which not only expands our understanding about ethylene signal regulation network in the carnation petal senescence process, but also provides potential targets with respect to breeding a cultivar of long-lived cut carnation.

Genome-wide characterization of DcHsp90 gene family in carnation (Dianthus caryophyllus L.) and functional analysis of DcHsp90-6 in heat tolerance.

Plant heat shock protein 90 (Hsp90) participates in various physiological processes including protein folding, degradation, and signal transduction. However, the DcHsp90 gene family in carnation (Dianthus caryophyllus L.) has not been systematically analyzed. We thoroughly examined and comprehensively analyzed the carnation DcHsp90 gene family in this study and discovered 9 DcHsp90 genes. Based on the phylogenetic examination, DcHsp90 proteins may be divided into two groups. DcHsp90 structural features were similar but varied between groups. Promoter analysis revealed the presence of many cis-acting elements, most of which were connected to growth and development, hormones, and stress. DcHsp90 genes may play distinct functions in heat stress response, according to gene expression analyses. The DcHsp90-6 was isolated, and its role in the reaction to heat stress was studied. Thermotolerance and superoxide dismutase activity in transgenic seedlings were enhanced by Arabidopsis overexpression of DcHsp90-6. After heat stress, transgenic plants' electrolyte leakage and malondialdehyde levels were much lower than wild-type plants. Furthermore, overexpression of DcHsp90-6 altered the expressions of stress-responsive genes such as AtHsp101, AtHsp90, AtGolS1, AtRS4/5, and AtHsfB1. This study provides comprehensive information on the DcHsp90 gene family and suggests that overexpressed DcHsp90-6 positively regulates thermotolerance highlighting the adaptation mechanism of carnation under heat stress.

DcWRKY33 promotes petal senescence in carnation (Dianthus caryophyllus L.) by activating genes involved in the biosynthesis of ethylene and abscisic acid and accumulation of reactive oxygen species.

Carnation (Dianthus caryophyllus L.) is one of the most famous and ethylene-sensitive cut flowers worldwide, but how ethylene interacts with other plant hormones and factors to regulate petal senescence in carnation is largely unknown. Here we found that a gene encoding WRKY family transcription factor, DcWRKY33, was significantly upregulated upon ethylene treatment. Silencing and overexpression of DcWRKY33 could delay and accelerate the senescence of carnation petals, respectively. Abscisic acid (ABA) and H2 O2 treatments could also accelerate the senescence of carnation petals by inducing the expression of DcWRKY33. Further, DcWRKY33 can bind directly to the promoters of ethylene biosynthesis genes (DcACS1 and DcACO1), ABA biosynthesis genes (DcNCED2 and DcNCED5), and the reactive oxygen species (ROS) generation gene DcRBOHB to activate their expression. Lastly, relationships are existed between ethylene, ABA and ROS. This study elucidated that DcWRKY33 promotes petal senescence by activating genes involved in the biosynthesis of ethylene and ABA and accumulation of ROS in carnation, supporting the development of new strategies to prolong the vase life of cut carnation.

The transcription factors DcHB30 and DcWRKY75 antagonistically regulate ethylene-induced petal senescence in carnation (Dianthus caryophyllus).

Although numerous transcription factors with antagonistic activities have been shown to contribute to growth and development, whether and how they regulate senescence in plants is largely unknown. In this study, we investigated the role of antagonistic transcription factors in petal senescence in carnation (Dianthus caryophyllus), one of the most common types of ethylene-sensitive cut flowers produced worldwide. We identified DcHB30 that encodes a ZF-HD transcription factor that is down-regulated in ethylene-treated petal transcriptomes. We found that silencing DcHB30 accelerated ethylene-induced petal senescence and that DcHB30 physically interacts with DcWRKY75, a positive regulator of ethylene-induced petal senescence. Phenotypic characterization and molecular evidence indicated that DcHB30 and DcWRKY75 competitively regulate the expression of their co-targeted genes DcACS1, DcACO1, DcSAG12, and DcSAG29 by reciprocally inhibiting the DNA-binding activity of each other on the gene promoters. This transcriptional regulation mechanism demonstrates that these transcription factors serve as positive and negative regulators in ethylene-induced petal senescence in carnation. Thus, our study provides insights into how antagonizing transcription factors regulate plant senescence.

Effect of natural products use prior to infection with COVID-19 on disease severity and hospitalization: A self-reported cross-sectional survey study.

Background: Managing coronavirus disease 2019 (COVID-19) using available resources is essential to reduce the health burden of disease. The severity of COVID-19 is affected by nutritional status. In this study the effect of natural product use prior to infection with COVID-19 on disease severity and hospitalization was explored. Methods: This was a cross-sectional study. Between March and July 2021, a self-administered survey was conducted in Jordan. Individuals who recovered from COVID-19 and were ≥18 years old were the study population. Study measures included the use of natural products, COVID-19 severity, and hospitalization status. A multivariate regression model was used for statistical analysis. Results: The mean age (mean ± SD) of the study sample (n=2,148) was 40.25 ± 15.58 years old. Multivariate logistic regression showed that the regular intake of carnation (OR [0.56], CI [0.37-0.85]), onion (OR [0.69], CI [0.52-0.92]), lemon (OR [0.68], CI [0.51-0.90]), and citrus fruits (OR [0.66], CI [0.50-0.89]) before infection were associated with a substantial reduction in COVID-19 severity (P<0.01). Also, the consumption of carnation (OR [0.55], CI [0.34-0.88]), lemon (OR [0.57], CI [0.42-0.78]), and citrus fruits (OR [0.61], CI [0.44-0.84]) were associated with a significant decrease in the frequency of COVID-19-induced hospitalization (P<0.01). Conclusions: Regular consumption of carnation, lemon, and citrus fruits before infection was associated with better outcomes for COVID-19. Studies on other populations are required to confirm these findings.

Identification of the DcHsp20 gene family in carnation (Dianthus caryophyllus) and functional characterization of DcHsp17.8 in heat tolerance.

MAIN CONCLUSION: 33 heat shock protein 20 (Hsp20) genes were identified from the carnation genome whose expression were altered by abiotic stresses. DcHsp17.8 may function to improve the heat resistance of Arabidopsis. Heat shock proteins 20 (Hsp20s) mainly function as molecular chaperones that play crucial roles in relieving abiotic stresses such as heat stress. In this study, we identified and characterized 33 DcHsp20 genes from the carnation genome that were classified into 9 subfamilies. Gene structure analysis showed that 25 DcHsp20 genes contained 1 intron whilst the remaining 8 DcHsp20 genes did not contain introns. Motif analysis found that DcHsp20 proteins were relatively conserved. Cis-regulatory elements analysis of the Hsp20 promoters revealed a number of cis-regulatory elements that regulate growth and development, hormone and stress responses. Gene expression analysis revealed that DcHsp20 genes had multiple response patterns to heat stress. The largest range of induction occurred in DcHsp17.8 after 1 h of heat stress. Under cold stress, or treatment with saline or abscisic acid, the expression of most DcHsp20 genes was inhibited. To further understand the function of DcHsp20 genes in response to heat stress, we overexpressed DcHsp17.8 in Arabidopis and the plants showed improved heat tolerance, O2- and H2O2 activities and photosynthetic capacity with reduced relative electrolyte leakage and malondialdehyde content. Gene expression analysis revealed that DcHsp17.8 modulated the expression of genes involved in antioxidant enzyme synthesis. Our data provided a solid foundation for the further detailed study of DcHsp20 genes.

DcWRKY75 promotes ethylene induced petal senescence in carnation (Dianthus caryophyllus L.).

Carnation (Dianthus caryophyllus L.) is one of the most important and typical ethylene sensitive cut flowers worldwide, although how ethylene influences the petal senescence process in carnation remains largely unknown. Here, we screened out one of the key transcription factors, DcWRKY75, using a constructed ethylene induced petal senescence transcriptome in carnation and found that it shows quick induction by ethylene treatment. Silencing of DcWRKY75 delays ethylene induced petal senescence in carnation. Molecular evidence confirms that DcWRKY75 can bind to the promoter regions of two main ethylene biosynthetic genes (DcACS1 and DcACO1) and a couple of senescence associated genes (DcSAG12 and DcSAG29) to activate their expression. Furthermore, we show that DcWRKY75 is a direct target gene of DcEIL3-1, which is a homolog of the ethylene signaling core transcription factor EIN3 in Arabidopsis. DcEIL3-1 can physically interact with DcWRKY75 and silencing of DcEIL3-1 also delays ethylene induced petal senescence in carnation and inhibits the ethylene induced expression of DcWRKY75 and its target genes. The present study demonstrates that the transcriptional regulation network is vitally important for ethylene induced petal senescence process in carnation and potentially in other ethylene sensitive cut flowers.

Flavonoid biosynthesis in Dianthus caryophyllus L. is early regulated during interaction with Fusarium oxysporum f. sp. dianthi.

Rooted cuttings from two carnation (Dianthus caryophyllus L.) cultivars showing contrasting responses to the vascular wilt caused by Fusarium oxysporum f. sp. dianthi (Fod) were inoculated with this phytopathogen, and some of the biochemical responses associated with flavonoid biosynthesis were investigated in the roots. The resistant cultivar ('Golem') showed a significant increase in the levels of phenolic and flavonoid compounds at 48-96 h post-inoculation (hpi) (α = 0.05). LC-MS-based analysis indicated that the flavonoids mainly included flavanol-type glycosides, especially quercetin and kaempferol aglycones. Quantification of the mRNA levels of genes encoding CHS (Chalcone Synthase), CHI (Chalcone Isomerase), FLS (Flavonol Synthase), and the transcription factor MYB11 by using reverse transcription quantitative polymerase chain reaction (RT-qPCR) indicated that the resistant cultivar exhibited higher expression levels of these genes and, therefore, showed more flavonoid accumulation at 96 hpi. The differences in the temporal regulation of the assessed variables during infection support the idea that the early expression of enzymes of the flavonoid biosynthesis pathway in carnation roots is linked to a resistance response to the hemibiotrophic pathogen Fod race 2. The present experimental approach is the first report describing the molecular mechanisms underlying flavonoid biosynthesis in carnation roots during their interaction with Fod.