Search Results (131)

Carotenoids are essential nutritional compounds in tomato (Solanum lycopersicum), and mutations in carotenoid biosynthesis genes can alter both fruit color and other quality traits. Carotenoid isomerase (CRTISO) converts prolycopene to all-trans-lycopene; its mutation results in the orange-fruited tangerine mutant characterized by prolycopene accumulation. However, the broader impacts of distinct SlCRTISO mutations on fruit quality traits and agronomic performance remain largely unexplored. Here, we evaluated agronomic traits and nutritional quality in two tangerine mutants carrying distinct SlCRTISO alleles. The yellow/orange-fruited tomato 5 (yft5) mutant, which harbors a single point mutation in SlCRTISO, exhibited lower flavonoids, chlorogenic acid, and prolycopene than the CRISPR/Cas9-mediated knockout mutants, but maintained fruit size and weight comparable to wild type, whereas knockout lines showed severely reduced fruit size, highlighting the potential agronomic advantage of yft5. Pearson correlation analysis revealed negative associations between abscisic acid (ABA) levels and rutin, chlorogenic acid, as well as the prolycopene-to-lycopene ratio during fruit ripening. Transcriptome analysis demonstrated that knockout mutations more profoundly affected primary metabolism than the yft5 point mutation. Notably, 9-cis-epoxycarotenoid dioxygenase 2 (NCED2), a gene involved in ABA biosynthesis, was specifically upregulated in yft5 fruits at the mature green stage (38 days post-anthesis (dpa)), consistent with ABA levels. Quality-related genes including chalcone synthase (CHS), citrate synthase (CS), and invertase (INV) were significantly upregulated in knockout lines during ripening. These findings reveal a trade-off between phenolic content and fruit size in tangerine tomatoes carrying distinct SlCRTISO alleles, positioning yft5 as a valuable genetic resource for breeding high-quality varieties. Full article

Emilia sonchifolia (L.) DC is a medicinal and edible herb of Asteraceae with Lingnan characteristics. Flavonoids are its core pharmacodynamic substances, but the molecular regulation mechanism of differential accumulation of flavonoids in different organs of this species is still unclear. In this study, the molecular basis of tissue-specific synthesis of flavonoids was analyzed by integrating UPLC-MS broad-target metabolome and Illumina high-throughput transcriptome with four tissues of Emilia sonchifolia: root, stem, leaf, and flower. The results showed that a total of 73 flavonoid metabolites were identified in the metabolome, including naringenin chalcone, luteolin, quercitrin, and other pharmacologically active substances. Multi-omics joint analysis showed that the floral organ was the core tissue for the synthesis and enrichment of flavonoids, and there were specific characteristic flavonoid subtypes in different tissues. A total of 211 differentially expressed genes related to the flavonoid synthesis pathway were screened by transcriptome analysis, including 16 flavonol synthases, five cinnamic acid 4-hydroxylases, and five chalcone synthases. The WGCNA and gene–metabolite association network showed that the transcription levels of key enzyme genes such as CHS, C4H, F3′H, and F3H were highly positively correlated with the accumulation of downstream flavonols. The qRT-PCR quantitative verification showed that the expression patterns of CHS1, CHI4, F3′H5, F3H, FLS4, and GT in the four tissues were highly consistent with the transcriptome sequencing results, which confirmed that the transcriptome data were reliable. For the first time, this study revealed the molecular regulatory network of tissue-specific accumulation of flavonoids in Emilia sonchifolia, and clarified that the flower organ was the optimal medicinal harvesting site of flavonoids. It provided key theoretical support for the breeding of high-efficacy Emilia sonchifolia germplasm, the development of flavonoid active ingredients, and the study of secondary metabolic evolution of Compositae plants. Full article

Black soybean is an important functional crop valued for its high anthocyanin content and associated health benefits, attracting increasing interest in plant breeding and mutation-based approaches to enhance its nutritional and agronomic traits. Here, we investigated a DT26 black seed-coat mutant (DT26BS) derived from the Vietnamese cultivar DT26 following gamma irradiation. Genetic analysis of F₂ populations indicated that the phenotype is controlled by a single recessive mutation, with additional epistatic interactions observed in other genetic backgrounds. MutMap analysis based on whole-genome sequencing of pooled F₂ individuals identified a candidate region on chromosome 8 corresponding to the I locus. A large deletion (~176 kb) was identified in this region, affecting multiple Chalcone Synthase (CHS) gene repeats, and which may disrupt RNAi-mediated silencing of CHS, thereby triggering anthocyanin restoration in the seed coat. PCR-based markers confirmed tight linkage between this deletion and the black seed-coat phenotype. Crosses with elite lines and further selection until F7 generations showed that the mutation has no adverse effects on major agronomic traits and is useful as a donor for developing improved lines with black seed coat, shorter maturity, high yield, and enhanced anthocyanin content for nutritional improvement. These results demonstrate the utility of MutMap for detecting irradiation-induced structural variants and extend its application to a non-reference elite tropical soybean background, providing a useful genetic resource for black soybean breeding. Full article

The chalcone synthase (CHS) genes play an important role in the biosynthesis of flavonoids and anthocyanins. However, research into the genomic characteristics and evolutionary patterns of the CHS gene family in lychee remains limited. In this study, six LcCHS genes were identified from the litchi genome. Most members of the LcCHS family consist of two exons and possess eight conserved motifs; furthermore, all members exhibit highly similar three-dimensional protein structures. LcCHS genes were closely related to homologs of Sapindaceae species and experienced gene duplication events during dicotevolution. Comparative colinearity analysis revealed conserved colinearity relationships among most LcCHS genes, and all members of the LcCHS family exhibited distinct colinearity with homologues in longan. LcCHS genes exhibited significant tissue-specific expression preferentially in reproductive tissues rather than vegetative tissues. Transcription analysis of ‘Jinggang Hongnuo’ litchi fruit development divided LcCHS genes into two negatively correlated expression modules and LcCHS members interacted with numerous transcription factors (e.g., flavonoid biosynthesis, hormone signaling and developmental regulation). Spatiotemporal expression profiling of LcCHS genes in ‘Guiwei’, ‘Feizixiao’ and ‘Ziniangxi’ litchi cultivars revealed obvious tissue specificity and developmental stage preference of these genes. Gene expression of LcCHS genes in ‘Guiwei’ high correlated with the flavonoid content. Furthermore, promoter cis-element analysis identified 34 types of cis elements in LcCHS promoters that participate in plant growth and development, hormone response and stress adaptation, implying that LcCHS genes may participate in litchi growth, hormone signal transduction, and stress-response processes. Comprehensive characterization of the chalcone synthase gene family could provide insights into the genetic improvement of litchi. Full article

Flavonoids are essential secondary metabolites that predominantly affect flower pigmentation in plants. Understanding the molecular mechanisms underlying flower color divergence is crucial for ornamental plant breeding. This study aimed to elucidate the factors responsible for the differences in color between white-petaled Cosmos bipinnatus and orange-petaled Cosmos sulphureus. We employed an integrated approach combining untargeted LC–MS/MS metabolomics and high-throughput transcriptome sequencing of fresh petals to analyze pigment composition and differential gene expression. Petal pigment extraction, total flavonoid quantification, and metabolomic profiling consistently revealed that differences in flavonoid abundance are responsible for flower color divergence between the two species. In contrast, carotenoids, previously considered potential contributors to flower coloration, were neither evident in the oil phase of the pigment extracts nor detected by metabolomic analysis. Flavonoid compounds accumulated at relatively high levels in the orange petals of C. sulphureus, reaching 11.36 times that of C. bipinnatus, contributing to its bright appearance. Transcriptomic analysis revealed differences in gene expression patterns between the two species, highlighting key candidate genes involved in the flavonoid biosynthesis pathway, such as chalcone synthase. These findings indicate that the orange coloration of C. sulphureus may be associated with CHS-regulated accumulation of naringenin chalcone and downstream compounds in the flavonoid metabolic pathway after CHS, providing valuable theoretical support for a deeper understanding of the causes underlying the differences in flower color between C. bipinnatus and the orange-petaled C. sulphureus. Full article

Peach (Prunus persica) fruit pigmentation is largely associated with anthocyanin accumulation, whereas colorless flavonols such as kaempferol glycosides may reflect alternative use of shared flavonoid precursors. To examine the relationship between anthocyanin and selected kaempferol glycoside accumulation, we analyzed 15 peach accessions classified by red, white, or yellow flesh pigmentation. Skin color was quantified using the a*/b* ratio, where a* represents redness/greenness and b* represents yellowness/blueness. Red-fleshed accessions showed higher skin a*/b* values and accumulated higher levels of total anthocyanins, particularly cyanidin-3-glucoside, than white and yellow accessions. In contrast, kaempferol-3-rhamnoside preferentially accumulated in white-fleshed accessions. Expression analysis of flavonoid pathway genes showed that dihydroflavonol 4-reductase (PpDFR) was more highly expressed in red accessions, whereas flavonol synthase (PpFLS) was more highly expressed in white accessions; chalcone synthase (PpCHS), flavanone 3-hydroxylase (PpF3H), flavonoid 3′-hydroxylase (PpF3′H), and anthocyanidin synthase (PpANS) showed no significant differences among color groups. Heterologous overexpression of PpF3′H in Arabidopsis thaliana, a well-characterized model plant for flavonoid biosynthesis, was associated with increased seed anthocyanin accumulation and a lower kaempferol-to-quercetin ratio, supporting its catalytic capacity to influence flavonoid composition in an exogenous system. Overall, these results indicate that differential anthocyanin and selected kaempferol glycoside accumulation in peach is associated with the relative expression patterns of branch-related flavonoid genes, particularly PpDFR and PpFLS. This study provides targeted metabolic and transcriptional evidence for understanding peach flesh and skin pigmentation and provides mechanistic insight into flavonoid branch competition linking gene expression patterns with metabolite allocation, and identifies candidate genes for improving fruit color and flavonoid-related nutritional quality. Full article

Safflower (Carthamus tinctorius L.) is an important medicinal plant widely used as a source of natural pigments. Flower color is a key trait affecting both ornamental and commercial value; however, the roles of long non-coding RNAs (lncRNAs) in safflower flower coloration remain largely unclear. In this study, strand-specific RNA sequencing was performed on three safflower varieties with distinct flower colors at different floral developmental stages. A total of 4851 lncRNAs were identified, including 940 natural antisense transcript (NAT) pairs. Among them, lncRNA MSTRG.28365 was identified as a natural antisense transcript paired with CtCHS.7, a chalcone synthase-like gene potentially involved in flavonoid biosynthesis. Expression analysis revealed that CtCHS.7 was highly expressed in the red-flowered variety, whereas MSTRG.28365 exhibited an opposite expression pattern, suggesting a potential regulatory association. Co-expression analysis further indicated that CtCHS.7 was associated with genes putatively involved in flavonoid modification, including UDP-glycosyltransferases and cytochrome P450 enzymes. Functional assays showed that the recombinant CtCHS.7 protein could catalyze the production of downstream flavonoid-related metabolites, although the detected product differed from canonical naringenin chalcone. These findings suggest that lncRNAs may participate in flower color variation and flavonoid biosynthesis-related processes in safflower. This study provides candidate regulatory elements for future functional validation of safflower flower coloration mechanisms. Full article

Spatholobus suberectus is a traditional edible, ornamental, and medicinal vine with an abundant flavonoid content in its dried stems. An integrated metabolomic and transcriptomic analysis was conducted on its vegetative organs to identify flavonoid biosynthesis pathways. The results showed that: (1) a total of 268 flavonoids could be identified among all the vegetative organs, and stems accumulated most of them; (2) a total of 449,569,220 clean reads were retained from all the vegetative organs (many genes related to key enzymes of flavonoid biosynthesis and transcription factors were found in all the compared groups between different organs; among them, 77 genes showed high correlations with key flavonoids in different pathways); and (3) PAL (encoding phenylalanine ammonia-lyase), 4CL (encoding 4-coumarate: CoA ligase), C4H (encoding cinnamate-4-hydroxylase), CHS (encoding chalcone synthase), DFR (encoding dihydroflavonol 4-reductase) and others showed positive correlations with L-phenylalanine and various flavonoids, while FLS (encoding flavonol synthase) showed negative correlations with many flavonols and flavonoids, as revealed through integrated analysis. This study reveals the flavonoid biosynthesis characteristics in S. suberectus and provides new insights into the utilization of its biological resources. Full article

Exogenously induced RNA interference (exoRNAi) is a powerful biotechnology tool for precise gene regulation. The plant chalcone synthase (CHS) gene serves as a valuable model for molecular biology due to its central role in flavonoid biosynthesis. However, there are currently very few studies addressing the advantages and disadvantages of in vitro (enzymatic) or in vivo (bacterial) methods for producing double-stranded RNA (dsRNA) for exogenous application. This study aims to optimize and compare the two methods for producing dsRNAs targeting the Arabidopsis thaliana CHS gene: enzymatic synthesis in vitro using a commercial kit and bacterial synthesis in vivo using an engineered E. coli HT115 (DE3) system. Bacterial synthesis conditions were optimized with respect to IPTG concentration and cultivation time, and the resulting dsRNA preparations were purified and quality-controlled. Their biological activities were assessed by treating A. thaliana plants and analyzing the effects on AtCHS gene expression and flavonoid production using qRT-PCR and HPLC-MS. The results demonstrated that purified AtCHS-dsRNA from both methods effectively suppressed AtCHS expression and downstream flavonoid biosynthetic gene expression, leading to significant reductions in anthocyanins and flavanols. This study confirmed the efficacy of exogenous dsRNAs in regulating plant metabolic pathways and provided a comparative analysis of dsRNA synthesis methods, highlighting their benefits and limitations for practical applications in plant biology and protection. Full article

Plasmid copy number (PCN) is a key factor limiting the expression level of heterologous proteins in yeast. Static strategies for enhancing PCN, such as reducing the transcriptional intensity of selection markers or increasing selection pressure, only maintain PCN at a single fixed level and struggle to achieve dynamic, precise, and reversible copy number regulation. This study established a dynamic plasmid copy number regulation strategy based on CRISPR interference (CRISPRi). Flexible control of PCN was achieved by designing specific guide RNAs (gRNAs) and integrating them into the inducible CRISPRi system. Optimization of the gRNA target site, inducer concentration, and induction timing resulted in a >2-fold increase in the fluorescence intensity of yeast-enhanced green fluorescent protein (yeGFP) compared with the group without induction. Using naringenin synthesis as proof-of-concept, this regulatory tool was applied to modulate the expression of chalcone synthase (CHS), the rate-limiting enzyme in naringenin biosynthesis. Finally, the yield of naringenin increased by 35.62% under the optimal induction conditions. Full article

Morus macroura ‘Panzhihua No. 1’ is a dual-purpose cultivar valued for its edible leaves and fruits. Derived from an ancient mulberry tree, it is a unique local germplasm resource. During asexual propagation, M. macroura exhibits sexual variation closely associated with fruit and leaf yield. To explore the molecular mechanisms of sexual dimorphism and its effects on nutritional traits, we integrated transcriptomic and metabolomic analyses of male and female inflorescences and leaves. Key sex-biased genes were enriched in plant hormone signaling, flavonoid biosynthesis, and carbohydrate metabolism pathways. Female plants had elevated expression of ethylene-responsive transcription factor 1 (ERF1), EIN3-binding F-box proteins (EBF1/2), and Chalcone synthase (CHS) genes and higher levels of bioactive flavonoids, including isoquercitrin and kaempferol derivatives. In contrast, male plants had increased expression of gibberellin 20-oxidase (GA20ox) and DELLA genes and accumulated glycosides, which are beneficial for leaf development. These findings reveal how sex-linked metabolic networks shape mulberry tissue functional profiles, providing molecular targets for breeding. Full article

Cibotium barometz (L.) J. Sm., a medicinally significant fern in traditional Chinese medicine, is little explored at the genomic level regarding its bioactive compounds. Using an integrated approach combining Illumina and PacBio sequencing technologies, we profiled its root, rachis, and pinna transcriptomes, identifying 12,718, 21,341, and 11,441 unigenes, respectively. Our analysis systematically characterized the transcriptional features of transcription factors (TFs), simple sequence repeats (SSRs), long non-coding RNAs (lncRNAs), and differentially expressed genes (DEGs). Enrichment analyses highlighted the roles of highly expressed unigenes in secondary metabolism. Seventeen key enzymes involved in polysaccharide biosynthesis showed tissue-specific expression patterns. Notably, total polysaccharide content correlated positively with UDP-arabinose 4-epimerase (UXE) expression but negatively with phosphoglucomutase (PGM) and 3,5-epimerase/4-reductase (UER1). Flavonoid accumulation inversely correlated with chalcone synthase (CHS) expression. Two lignin pathways (H-lignin and G-lignin) were characterized, with phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H), and cinnamyl alcohol dehydrogenase (CAD) as key genes. The absence of ferulate-5-hydroxylase (F5H) explains the undetected S-lignin pathway. Regulatory network analysis revealed positive correlations between PAL expression and NAC72/NAC78/WRKY35 and C4H expression and WRKY65/WRKY69/WRKY71, while a negative correlation was revealed between flavonoid 3′,5′-hydroxylase (F3′5′H) and MYB3R4. This study provides comprehensive transcriptomic insights into C. barometz bioactive compound biosynthesis, serving as a foundation for mechanistic research. Full article

Aquilegia (Ranunculaceae), a genus of perennial herbs characterized by elegant leaves and unique flowers, exhibits promising application prospects in Northeast China. Chalcone synthase participates in the first enzymatic reaction of the anthocyanin biosynthesis pathway and plays a crucial role in flower color formation. In the present study, eight AoCHSs were identified and a comprehensive analysis of AoCHSs was then carried out, considering physical and chemical properties, conservative motifs, phylogenetic relationships, and cis-acting elements. The expression patterns of the AoCHSs were analyzed based on the transcriptome of A. oxysepala, which differs in sepal color between two species, to identify the candidate genes involved in flower color variation. AoCHS2/3/5 expression levels were found to be upregulated at PrA stage in A. oxysepala with dark purple sepals, while remaining consistently low in the pale-yellow species. Combining KEGG annotations and expression patterns, AoCHS5 was identified as a key gene for anthocyanin biosynthesis in Aquilegia that could play a role in the variation in flower color. The results of correlation network analysis showed that AoCHS5 was highly associated with MYB, bHLH, and WRKY. These results provided genetic resources for accelerating the molecular breeding of innovative Aquilegia flower colors. Full article

Chalcone Synthase (CHS) plays a vital role in flavonoid synthesis, influencing plant growth, development, and responses to both biotic and abiotic stress. In this study, 11 CHS genes were identified in Poncirus trifoliata using bioinformatics methods, with their distribution across five chromosomes and unassigned contigs. Each gene contains 2–3 exons and 3–8 conserved motifs. In silico prediction suggested that the PtrCHS proteins are localized in the cytoplasm. PtrCHS9 and PtrCHS11 share identical protein tertiary structures. Phylogenetic analysis classified the CHS family members into four subgroups. Synteny analysis revealed one set of collinear gene pairs within Poncirus trifoliata. Between Poncirus trifoliata and Arabidopsis thaliana, two sets of collinear gene pairs were identified, while one such set was found between Poncirus trifoliata and Oryza sativa. Promoter element analysis showed the presence of various hormone response and stress response elements within PtrCHS promoters. RNA-Seq data demonstrated tissue-specific expression patterns of PtrCHSs. RT-qPCR results indicated that all CHS genes, except PtrCHS11, respond to salt stress with dynamic, member-specific patterns. Additionally, four PtrCHSs (PtrCHS3, PtrCHS5, PtrCHS7, and PtrCHS10) were significantly upregulated in response to cold treatment. Notably, PtrCHS7 and PtrCHS10 maintained high expression levels at both 6 and 12 h, implying they may be key players in cold stress response in Poncirus trifoliata. Clones of PtrCHS7 and PtrCHS10 were obtained, and overexpression vectors were constructed in preparation for gene transformation. Overall, this study provides a solid foundation for future research into the functions of the PtrCHSs. Full article

Flavonoids serve as crucial plant antioxidants in drought tolerance, yet their antioxidant regulatory mechanisms within mycorrhizal plants remain unclear. In this study, using a two-factor design, trifoliate orange (Poncirus trifoliata (L.) Raf.) seedlings in the four-to-five-leaf stage were either inoculated with Funneliformis mosseae or not, and subjected to well-watered (70–75% of field maximum water-holding capacity) or drought stress (50–55% field maximum water-holding capacity) conditions for 10 weeks. Plant growth performance, photosynthetic physiology, leaf flavonoid content and their antioxidant capacity, reactive oxygen species levels, and activities and gene expression of key flavonoid biosynthesis enzymes were analyzed. Although drought stress significantly reduced root colonization and soil hyphal length, inoculation with F. mosseae consistently enhanced the biomass of leaves, stems, and roots, as well as root surface area and diameter, irrespective of soil moisture. Despite drought suppressing photosynthesis in mycorrhizal plants, F. mosseae substantially improved photosynthetic capacity (measured via gas exchange) and optimized photochemical efficiency (assessed by chlorophyll fluorescence) while reducing non-photochemical quenching (heat dissipation). Inoculation with F. mosseae elevated the total flavonoid content in leaves by 46.67% (well-watered) and 14.04% (drought), accompanied by significantly enhanced activities of key synthases such as phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), 4-coumarate:coA ligase (4CL), and cinnamate 4-hydroxylase (C4H), with increases ranging from 16.90 to 117.42% under drought. Quantitative real-time PCR revealed that both mycorrhization and drought upregulated the expression of PtPAL1, PtCHI, and Pt4CL genes, with soil moisture critically modulating mycorrhizal regulatory effects. In vitro assays showed that flavonoid extracts scavenged radicals at rates of 30.07–41.60% in hydroxyl radical (•OH), 71.89–78.06% in superoxide radical anion (O₂^•−), and 49.97–74.75% in 2,2-diphenyl-1-picrylhydrazyl (DPPH). Mycorrhizal symbiosis enhanced the antioxidant capacity of flavonoids, resulting in higher scavenging rates of •OH (19.07%), O₂^•− (5.00%), and DPPH (31.81%) under drought. Inoculated plants displayed reduced hydrogen peroxide (19.77%), O₂^•− (23.90%), and malondialdehyde (17.36%) levels. This study concludes that mycorrhizae promote the level of total flavonoids in trifoliate orange by accelerating the flavonoid biosynthesis pathway, hence reducing oxidative damage under drought. Full article