Transcriptome wide analysis of MADS box genes in Crocus sativus and interplay of CstMADS19-CstMADS26 in orchestrating apocarotenoid biosynthesis.

Flowers of Crocus sativus L. are immensely important not only for arrangement of floral whorls but more because each floral organ is dominated by a different class of specialized compounds. Dried stigmas of C. sativus flowers form commercial saffron, and are known to accumulate unique apocarotenoids like crocin, picrocrocin and safranal. Inspite of being a high value crop, the molecular mechanism regulating flower development in Crocus remains largely unknown. Moreover, it would be very interesting to explore any co-regulatory mechanism which controls floral architecture and secondary metabolic pathways which exist in specific floral organs. Here we report transcriptome wide identification of MADS box genes in Crocus. A total of 39 full length MADS box genes were identified among which three belonged to type I and 36 to type II class. Phylogeny classified them into 11 sub-clusters. Expression pattern revealed some stigma up-regulated genes among which CstMADS19 encoding an AGAMOUS gene showed high expression. Transient over-expression of CstMADS19 in stigmas of Crocus resulted in increased crocin by enhancing expression of pathway genes. Yeast one hybrid assay demonstrated that CstMADS19 binds to promoters of phytoene synthase and carotenoid cleavage dioxygenase 2 genes. Yeast two hybrid and BiFC assays confirmed interaction of CstMADS19 with CstMADS26 which codes for a SEPALATA gene. Co-overexpression of CstMADS19 and CstMADS26 in Crocus stigmas enhanced crocin content more than was observed when genes were expressed individually. Collectively, these findings indicate that CstMADS19 functions as a positive regulator of stigma based apocarotenoid biosynthesis in Crocus.

Metabolomics reveals a key role of salicylic acid in embryo abortion underlying interspecific hybridization between Hydrangea macrophylla and H. arborescens.

KEY MESSAGE: Embryo abortion at the heart-shaped stage is the main reason for the failure of interspecific hybridization of hydrangea, and salicylic acid plays a key role during embryo abortion. Difficulties in obtaining seeds from interspecific hybridization between Hydrangea macrophylla and H. arborescens had severely restricted the process of breeding new hydrangea varieties. To clarify the cause of reproductive barriers, an interspecific hybridization was made between H. macrophylla 'Endless Summer' (female parent) and H. arborescens 'Annabelle' (male parent). The results showed that both parents' floral organs developed normally, 'Annabelle' had high pollen viability (84.83% at 8 h after incubation), and the pollen tube could enter into the ovule of 'Endless Summer' at 72 h after pollination. Therefore, the pre-fertilization barrier was not the main reason for the failure of interspecific hybridization. However, observation of the embryo development by paraffin sections showed that the embryo was aborted at the heart-shaped stage. In addition, salicylic acid (SA) content was significantly higher (fourfold, P < 0.01) at 21 days after pollination (DAP) as compared to that of 17 DAP, which means SA may be closely correlated with embryo development. A total of 957 metabolites were detected, among which 78 were significantly different. During the embryo abortion, phenylpropanoids and polyketides were significantly down-regulated, while organic oxygen compounds were significantly up-regulated. Further analysis indicated that the metabolic pathway was enriched in the shikimic acid biosynthesis pathway, which suggests that more SA was synthesized. Taken together, it can be reasonably speculated that SA plays a key role leading to embryo abortion underlying the interspecific hybridization between Hydrangea macrophylla and H. arborescens. The result is helpful to direct the breeding of hydrangea through distant hybridization.

A novel strategy to map a locus associated with flowering time in canola (Brassica napus L.).

Flowering time is an important agronomic trait for canola breeders, as it provides growers with options for minimizing exposure to heat stress during flowering and to more effectively utilize soil moisture. Plants have evolved various systems to control seasonal rhythms in reproductive phenology including an internal circadian clock that responds to environmental signals. In this study, we used canola cultivar 'Westar' as a recurrent parent and canola cultivar 'Surpass 400' as the donor parent to generate a chromosome segment substitution line (CSSL) and to map a flowering time locus on chromosome A10 using molecular marker-assisted selection. This CSSL contains an introgressed 4.6 mega-bases (Mb) segment (between 13 and 17.6 Mb) of Surpass 400, which substantially delayed flowering compared with Westar. To map flowering time gene(s) within this locus, eight introgression lines (ILs) were developed carrying a series of different lengths of introgressed chromosome A10 segments using five co-dominant polymorphic markers located at 13.5, 14.0, 14.5, 15.0, 15.5, and 16.0 Mb. Eight ILs were crossed with Westar reciprocally and flowering time of resultant 16 F1 hybrids and parents were evaluated in a greenhouse (2021 and 2022). Four ILs (IL005, IL017, IL035, and IL013) showed delayed flowering compared to Westar (P < 0.0001), and their reciprocal crosses displayed a phenotype intermediate in flowering time of both homozygote parents. These results indicated that flowering time is partial or incomplete dominance, and the flowering time locus mapped within a 1 Mb region between two co-dominant polymorphic markers at 14.5-15.5 Mb on chromosome A10. The flowering time locus was delineated to be between 14.60 and 15.5 Mb based on genotypic data at the crossover site, and candidate genes within this region are associated with flowering time in canola and/or Arabidopsis. The co-dominant markers identified on chromosome A10 should be useful for marker assisted selection in breeding programs but will need to be validated to other breeding populations or germplasm accessions of canola.

AINTEGUMENTA-LIKE genes regulate reproductive growth and bud dormancy in Platanus acerifolia.

KEY MESSAGE: Platanus acerifolia AIL genes PaAIL5a/b and PaAIL6b participate in FT-AP1/FUL-AIL pathways to regulate bud dormancy. In addition, PaAIL6a/b can promote flowering, and PaAIL5b and PaAIL6b affect floral development. Bud dormancy and floral induction are essential processes for perennial plants, they are both regulated by photoperiod, temperature, and hormones, indicating the existence of common regulators for both processes. AINTEGUMENTA-LIKE (AIL) genes regulate reproductive growth of annual plants, including floral induction and flower development, and their homologs in poplar and grape act downstream of the florigen gene FT and the floral meristem identity genes AP1/FUL and function to maintain growth and thus inhibit dormancy induction. However, it is not known whether AIL homologs participate in the reproduction processes in perennials and whether the Platanus acerifolia AIL genes are involved in dormancy. P. acerifolia is a perennial woody plant whose reproductive growth is strongly associated with dormancy. Here, we isolated four AIL homologs from P. acerifolia, PaAIL5a, PaAIL5b, PaAIL6a, and PaAIL6b, and systematically investigated their functions by ectopic-overexpression in tobacco. The findings demonstrate that PaAIL5a/b and PaAIL6b respond to short day, low temperature, and hormone signals and act as the components of the FT-AP1/FUL-AIL pathway to regulate the bud dormancy in P. acerifolia. Notably, PaAIL5a/b and PaAIL6b function downstream of PaFTL-PaFUL1/2/3 to inhibit the dormancy induction and downstream of PaFT-PaFUL2/3 to promote the dormancy release. In addition, PaAIL6a/b were found to accelerate flowering in transgenic tobacco, whereas PaAIL5b and PaAIL6b affected the flower development. Together, our results suggest that PaAIL genes may act downstream of different PaFT/PaFTL and PaFUL proteins to fulfill conservative and diverse roles in floral initiation, floral development, and dormancy regulation in P. acerifolia.

Functional and transcriptional regulation of the anthocyanidin acyl modifier gene Gs5AT of Gentiana sino-ornata.

The Chinese gentian, Gentiana sino-ornata produces brilliant blue flowers. To investigate the biological function and transcriptional regulation mechanism of the anthocyanin 5-O-acyltransferase gene (Gs5AT ) in the corolla, it is beneficial to analyse the mechanism of blue flower colour presentation. In this investigation, we obtained the CDS and promoter sequences of the gene Gs5AT . Yeast one-hybrid experiments were used to identify the transcription factor GsbHLH7 that activates the gene Gs5AT . According to quantitive reverse transcription polymerase chain reaction analysis, the expression of the gene Gs5AT was significantly and positively correlated with the gene GsbHLH7 . The colour phenotype of the flowers was significantly altered by the virus-induced gene silencing transduction of Gs5AT and GsbHLH7 , with GsbHLH7 silencing producing more pronounced changes in the corolla colour than Gs5AT . The expression of GsF3'5'H , GsDFR , GsANS , Gs3GT , and Gs5GT all fell to varying degrees after GsbHLH7 silencing, indicating that GsbHLH7 may regulate transcription of these genes as well as Gs5AT . The results of this study indicate that Gs5AT was positively regulated by the GsbHLH7 , and thus affects the colour presentation of the blue corolla.

A multiscale approach to understanding the shared blue-orange flower color polymorphism in two Lysimachia species.

BACKGROUND: Polymorphisms are common in nature, but they are rarely shared among closely related species. Polymorphisms could originate through convergence, ancestral polymorphism, or introgression. Although shared neutral genomic variation across species is commonplace, few examples of shared functional traits exist. The blue-orange petal color polymorphisms in two closely related species, Lysimachia monelli and L. arvensis were investigated with UV-vis reflectance spectra, flavonoid biochemistry, and transcriptome comparisons followed by climate niche analysis. RESULTS: Similar color morphs between species have nearly identical reflectance spectra, flavonoid biochemistry, and ABP gene expression patterns. Transcriptome comparisons reveal two orange-specific genes directly involved in both blue-orange color polymorphisms: DFR-2 specificity redirects flux from the malvidin to the pelargonidin while BZ1-2 stabilizes the pelargonidin with glucose, producing the orange pelargonidin 3-glucoside. Moreover, a reduction of F3'5'H expression in orange petals also favors pelargonidin production. The climate niches for each color morph are the same between the two species for three temperature characteristics but differ for four precipitation variables. CONCLUSIONS: The similarities in reflectance spectra, biochemistry, and ABP genes suggest that a single shift from blue-to-orange shared by both lineages is the most plausible explanation. Our evidence suggests that this persistent flower color polymorphism may represent an ancestrally polymorphic trait that has transcended speciation, yet future analyses are necessary to confidently reject the alternative hypotheses.

Cytological analysis of flower development, insights into suitable growth area and genomic background: implications for Glehnia littoralis conservation and sustainable utilization.

BACKGROUND: Glehnia littoralis F. Schmidt ex Miq., an endangered plant species with significant medicinal, edible, and ecological value, is now a central concern for conservation and sustainable utilization. Investigating the physiological and ecological mechanisms leading to its endangerment and elucidating its genetic background constitutes the foundation for conducting in-depth research on G. littoralis. RESULTS: Our observations have revealed a significant degree of floral sterility in wild populations of G. littoralis. The inflorescences of G. littoralis are classified into three types: completely fertile, completely sterile, and partially fertile compound umbels. Moreover, the flowers of G. littoralis can be categorized into fertile and sterile types. Sterile flowers exhibited abnormalities in the stigma, ovary, and ovules. This study is the first to discover that the presence or absence of a giant cell at the funiculus during the initiation of ovule primordium determines whether the flower can develop normally, providing cytological evidence for female sterility in G. littoralis. Conversely, both fertile and sterile flowers produced normally developed pollen. Field observations have suggested that robust plants bear more fertile umbels, while weaker ones have fewer or even no fertile umbels, indicating a close relationship between flower fertility and plant nutritional status. Our model correctly predicted that the eastern coastal regions of China, as well as prospective areas in Neimenggu and Sichuan, are suitable environments for its cultivation. Additionally, Using flow cytometry and genome survey, we estimated the genome size of G. littoralis to be 3.06 Gb and the heterozygosity to be 4.58%. CONCLUSION: The observations and findings presented in this study were expected to provide valuable insights for further conserving its genetic resources and sustainable utilization of G. littoralis.

Expression of AtNF-YB1 activates early flowering, showing potential in breeding hybrid rice.

The nuclear factor Y (NF-Y) has been shown to be involved in plant growth and development in response to various environmental signals. However, the integration of these mechanisms into breeding practices for new cultivars has not been extensively investigated. In this study, the Arabidopsis gene AtNF-YB1 was introduced into rice, including inbred Kasalath and the hybrids Jinfeng × Chenghui 727 and Jinfeng × Chuanhui 907. The obtained transgenic rice showed early flowering under both natural long day (NLD) and natural short day (NSD) conditions. For the inbred Kasalath, the transgenic lines clearly showed a shorter plant height and lower grain yield, with a decrease in spike length and grain number but more productive panicles. However, the hybrids with AtNF-YB1 had much smaller or even zero reduction in spike length and grain number and more productive panicles. Thus, maintained or even increased grain yields of the transgenic hybrids were recorded under the NLD conditions. Quantitative PCR analysis indicated that the rice flowering initiation pathways were early activated via the suppression of Ghd7 induction in the transgenic rice. RNA-Seq further demonstrated that three pathways related to plant photosynthesis were markedly upregulated in both Jinfeng B and the hybrid Jinfeng × Chuanhui 907 with AtNF-YB1 expression. Moreover, physiological experiments showed an upregulation of photosynthetic rates in the transgenic lines. Taken together, this study suggests that AtNF-YB1 expression in rice not only induces early flowering but also benefits photosynthesis, which might be used to develop hybrid varieties with early ripening.

A Plant biostimulant prepared from Ascophyllum nodosum Induces Flowering by Regulating the MIR156-mediated Age Pathway in Arabidopsis.

Flowering, the change from vegetative development to the reproductive phase, represents a crucial and intricate stage in the life cycle of plants, which is tightly controlled by both internal and external factors. In this study, we investigated the effect of Ascophyllum nodosum extract (ANE) on the flowering time of Arabidopsis. We found that a 0.1% concentration of ANE induced flowering in Arabidopsis, accompanied by the upregulation of key flowering time genes: FT (FLOWERING LOCUS T), SOC1 (SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1), and LFY (LEAFY). Further investigation showed that ANE specifically promotes flowering through the MIR156-mediated age pathway. ANE treatment resulted in the repression of negative regulator genes, MIR156, while simultaneously enhancing the expression of positive regulator genes, including SPLs and MIR172. This, in turn, led to the downregulation of AP2-like genes, which are known as floral repressors. It is worth noting that ANE did not alleviate the late flowering phenotype of MIR156-overexpressing plants and spl mutants. Furthermore, ANE-derived fucoidan mimics the function of sugars in regulating MIR156, closely mirroring the effects induced by ANE treatments. It suppresses the transcript levels of MIR156 and AP2-like genes while inducing those of SPLs and MIR172, thereby reinforcing the involvement of fucoidan in the control of flowering by ANE. In summary, our results demonstrate that ANE induces flowering by modulating the MIR156-SPL module within the age pathway, and this effect is mediated by fucoidan.

CLAVATA3 Signaling Buffers Arabidopsis Shoot Apical Meristem Activity in Response to Photoperiod.

Land plants grow throughout their life cycle via the continuous activity of stem cell reservoirs contained within their apical meristems. The shoot apical meristem (SAM) of Arabidopsis and other land plants responds to a variety of environmental cues, yet little is known about the response of meristems to seasonal changes in day length, or photoperiod. Here, the vegetative and reproductive growth of Arabidopsis wild-type and clavata3 (clv3) plants in different photoperiod conditions was analyzed. It was found that SAM size in wild-type Arabidopsis plants grown in long-day (LD) conditions gradually increased from embryonic to reproductive development. clv3 plants produced significantly more leaves as well as larger inflorescence meristems and more floral buds than wild-type plants in LD and short-day (SD) conditions, demonstrating that CLV3 signaling limits vegetative and inflorescence meristem activity in both photoperiods. The clv3 phenotypes were more severe in SDs, indicating a greater requirement for CLV3 restriction of SAM function when the days are short. In contrast, clv3 floral meristem size and carpel number were unchanged between LD and SD conditions, which shows that the photoperiod does not affect the regulation of floral meristem activity through the CLV3 pathway. This study reveals that CLV3 signaling specifically restricts vegetative and inflorescence meristem activity in both LD and SD photoperiods but plays a more prominent role during short days.

Robust organ size in Arabidopsis is primarily governed by cell growth rather than cell division patterns.

Organ sizes and shapes are highly reproducible, or robust, within a species and individuals. Arabidopsis thaliana sepals, which are the leaf-like organs that enclose flower buds, have consistent size and shape, indicating robust development. Cell growth is locally heterogeneous due to intrinsic and extrinsic noise. To achieve robust organ shape, fluctuations in cell growth must average to an even growth rate, which requires that fluctuations are uncorrelated or anti-correlated in time and space. Here, we live image and quantify the development of sepals with an increased or decreased number of cell divisions (lgo mutant and LGO overexpression, respectively), a mutant with altered cell growth variability (ftsh4), and double mutants combining these. Changes in the number of cell divisions do not change the overall growth pattern. By contrast, in ftsh4 mutants, cell growth accumulates in patches of over- and undergrowth owing to correlations that impair averaging, resulting in increased organ shape variability. Thus, we demonstrate in vivo that the number of cell divisions does not affect averaging of cell growth, preserving robust organ morphogenesis, whereas correlated growth fluctuations impair averaging.

SlJMJ14, identified via QTL­seq and fine mapping, controls flowering time in tomatoes.

KEY MESSAGE: A major QTL, qLF2.1, for flowering time in tomatoes, was fine mapped to chromosome 2 within a 51.37-kb interval, and the SlJMJ14 gene was verified as the causal gene by knockout. Tomato flowering time is an important agronomic trait that affects yield, fruit quality, and environmental adaptation. In this study, the high-generation inbred line 19108 with a late-flowering phenotype was selected for the mapping of the gene that causes late flowering. In the F2 population derived from 19108 (late flowering) × MM (early flowering), we identified a major late-flowering time quantitative trait locus (QTL) using QTL-seq, designated qLF2.1. This QTL was fine mapped to a 51.37-kb genomic interval using recombinant analysis. Through functional analysis of homologous genes, Solyc02g082400 (SlJMJ14), encoding a histone demethylase, was determined to be the most promising candidate gene. Knocking out SlJMJ14 in MM resulted in a flowering time approximately 5-6 days later than that in the wild-type plants. These results suggest that mutational SlJMJ14 is the major QTL for the late-flowering phenotype of the 19108 parental line.

Identification and characterization of QSFS.sau-MC-5A for sterile florets genetically independent of fertile ones per spike in wheat.

KEY MESSAGE: A major and stable QTL for sterile florets per spike and sterile florets per spikelet was identified, it was mapped within a 2.22-Mb interval on chromosome 5AL, and the locus was validated using two segregating populations with different genetic backgrounds. Both the number of fertile florets per spike (FFS) and the number of sterile florets per spike (SFS) significantly influence the final yield of wheat (Triticum aestivum L.), and a trade-off theoretically exists between them. To enhance crop yield, wheat breeders have historically concentrated on easily measurable traits such as FFS, spikelets per spike, and spike length. Other traits of agronomic importance, including SFS and sterile florets per spikelet (SFPs), have been largely overlooked. In the study, reported here, genetic bases of SFS and SFPs were investigated based on the assessment of a population of recombinant inbred lines (RILs) population. The RIL population was developed by crossing a spontaneous mutant with higher SFS (msf) with the cultivar Chuannong 16. A total of 10 quantitative trait loci (QTL) were identified, with QSFS.sau-MC-5A for SFS and QSFPs.sau-MC-5A for SFPs being the major and stable ones, and they were co-located on the long arm of chromosome 5A. The locus was located within a 2.22-Mb interval, and it was further validated in two additional populations based on a tightly linked Kompetitive Allele-Specific PCR (KASP) marker, K_sau_5A_691403852. Expression differences and promoter sequence variations were observed between the parents for both TraesCS5A03G1247300 and TraesCS5A03G1250300. The locus of QSFS.sau-MC-5A/QSFPs.sau-MC-5A showed a significantly positive correlation with spike length, florets in the middle spikelet, and total florets per spike, but it showed no correlation with either kernel number per spike (KNS) or kernel weight per spike. Appropriate nitrogen fertilizer application led to reduced SFS and increased KNS, supporting results from previous reports on the positive effect of nitrogen fertilizer on wheat spike and floret development. Based on these results, we propose a promising approach for breeding wheat cultivars with multiple fertile florets per spike, which could increase the number of kernels per spike and potentially improve yield. Collectively, these findings will facilitate further fine mapping of QSFS.sau-MC-5A/QSFPs.sau-MC-5A and be instrumental in strategies to increase KNS, thereby enhancing wheat yield.

Integrative analysis of miRNA profile and degradome reveals post-transcription regulation involved in fragrance formation of Rosa rugosa.

Rosa rugosa is renowned for its fragrant essential oils (EOs) including the primary volatile compounds such as terpenes (geraniol and citronellol) and 2-phenylethanol. While the role of miRNAs in plant secondary metabolism has been explored, their involvement in EOs metabolism remains largely unknown. Sequencing of the petals of R. rugosa identified 383 conserved miRNAs and 625 novel miRNAs including 53 miRNAs differentially expressed in a strong fragrance variety R. rugosa 'White Purple Branch'. Degradome sequencing predicted 1969 targets enriched in GO terms involved in the negative regulation of macromolecule metabolic process. Furthermore, 122 targets of differentially expressed miRNAs were enriched in phenylalanine metabolism and other KEGG pathways. A post-transcriptional regulation network of 52 miRNAs and 70 miRNA-transcription factor modules target terpene and 2-phenylethanol biosynthesis pathways. Six interactions including miR535f-RrHMGR, NOV146-RrNUDX1, miR166l-RrHY5 and miR156c-RrSPL2 were validated using RNA ligase-mediated RACE. Sequence alignment revealed that the NOV146-RrNUDX1 was conserved in the Rosa genus. Moreover, weaker silencing of RrNUDX1 by NOV146 contributed to the stronger fragrance of R. rugosa. These findings offer a comprehensive understanding of the post-transcriptional regulation involved in essential oil biosynthesis and identify candidate miRNAs for further genetic improvement of EO yields in R. rugosa.

SUPERMAN genes: uncovering a new function in the development of complex inflorescences.

The Arabidopsis SUPERMAN (SUP) gene and its orthologs in eudicots are crucial in regulating the number of reproductive floral organs. In Medicago truncatula, in addition to this function, a novel role in controlling meristem activity during compound inflorescence development was assigned to the SUP-ortholog (MtSUP). These findings led us to investigate whether the role of SUP genes in inflorescence development was legume-specific or could be extended to other eudicots. To assess that, we used Solanum lycopersicum as a model system with a cymose complex inflorescence and Arabidopsis thaliana as the best-known example of simple inflorescence. We conducted a detailed comparative expression analysis of SlSUP and SUP from vegetative stages to flower transition. In addition, we performed an exhaustive phenotypic characterisation of two different slsup and sup mutants during the plant life cycle. Our findings reveal that SlSUP is required for precise regulation of the meristems that control shoot and inflorescence architecture in tomato. In contrast, in Arabidopsis, SUP performs no meristematic function, but we found a role of SUP in floral transition. Our findings suggest that the functional divergence of SUP-like genes contributed to the modification of inflorescence architecture during angiosperm evolution.

Integrated transcriptome and endogenous hormone analyses reveal the factors affecting the yield of Camellia oleifera.

Camellia oleifera is an important woody oil tree in China, in which the flowers and fruits appear during the same period. The endogenous hormone changes and transcription expression levels in different parts of the flower tissue (sepals, petals, stamens, and pistils), flower buds, leaves, and seeds of Changlin 23 high-yield (H), Changlin low-yield (L), and control (CK) C. oleifera groups were studied. The abscisic acid (ABA) content in the petals and stamens in the L group was significantly higher than that in the H and CK groups, which may be related to flower and fruit drops. The high N6-isopentenyladenine (iP) and indole acetic acid (IAA) contents in the flower buds may be associated with a high yield. Comparative transcriptome analysis showed that the protein phosphatase 2C (PP2C), jasmonate-zim-domain protein (JAZ), and WRKY-related differentially expressed genes (DEGs) may play an important role in determining leaf color. Gene set enrichment analysis (GSEA) comparison showed that jasmonic acid (JA) and cytokinin play an important role in determining the pistil of the H group. In this study, endogenous hormone and transcriptome analyses were carried out to identify the factors influencing the large yield difference in C. oleifera in the same year, which provides a theoretical basis for C. oleifera in the future.

Physiological and Transcriptome Analyses Reveal the Effects of Fertilization on the Yield of Winter Wheat and on the Photosynthetic Performance of Leaves during the Flowering Period.

Fertilization significantly affects the growth and development of wheat. However, the precise mechanisms underlying gene regulation during flowering in response to fertilization deficiency remain elusive. In this study, fertilization (F) and non-fertilization (CK) ) treatments were set up to reveal examine the effect of fertilization on the photosynthetic capacity of winter wheat during the flowering period through physiological, biochemical, and transcriptome analyses. Upon analyzing analysing their yield, leaf photosynthetic system exchange parameters during flowering, antioxidant enzyme activity, and endogenous hormone parameters, we found that the F treatment resulted in higher net photosynthetic rates during flowering periods than the CK treatment. The superoxide dismutase (SOD) (83.92%), peroxidase (POD) (150.75%), and catalase (CAT) (22.74%) activities of leaves in treated with F during the flowering period were notably elevated compared to those of CK-treated leaves. Abscisic acid (ABA) (1.86%) and gibberellin acid (GA3) (33.69%) levels were reduced, whereas Auxin auxin (IAA) (98.27%) content was increasedwas increased under F treatment compared to those the results under the CK treatment. The chlorophyll a (32.53%), chlorophyll b (56%), total chlorophyll (37.96%), and carotenoid contents (29.80%) under F treatment were also increased compared to CK., exceeded exceeding those obtained under the CK treatment. Furthermore, transcriptional differences between the F and CK conditions were analyzed, and key genes were screened and validated by using q-PCR. Transcriptome analysis identified 2281 differentially expressed genes (DEGs), with enriched pathways related to photosynthesis and light harvesting. DEGs were subjected to cluster simulation, which revealed that 53 DEGS, both up- and down-regulated, responded to the F treatment. qRT-PCR-based validation confirmed the differential expression of genes associated with carbohydrate transport and metabolism, lipid transport, and signal transduction. This study revealed distinctive transcriptional patterns and crucial gene regulation networks in wheat during flowering under fertilization, providing transcriptomic guidance for the precise regulation of wheat breeding.

Selection of Reference Genes of Flower Development in Ludisia discolor.

Background: RT-qPCR is a powerful strategy for recognizing the most appropriate reference genes, which can successfully minimize experimental mistakes through accurate normalization. Ludisia discolor, recognized for its ornamental value, features little, distinctive blossoms with twisted lips and gynostemium showing chiral asymmetry, together with striking blood-red fallen leaves periodically marked with golden blood vessels. Methods and Results: To ensure the accuracy of qRT-PCR, selecting appropriate reference genes for quantifying target gene expression levels is essential. This study aims to identify stable reference genes during the development of L. discolor. In this study, the entire floral buds, including the lips and gynostemium from different development stages, were taken as materials. Based upon the transcriptome information of L. discolor, nine housekeeping genes, ACT, HIS, EF1-α1, EF1-α2, PP2A, UBQ1, UBQ2, UBQ3, and TUB, were selected in this research study as prospect interior referral genes. The expression of these nine genes were found by RT-qPCR and afterwards comprehensively examined by four software options: geNorm, NormFinder, BestKeeper, and ΔCt. The outcomes of the analysis showed that ACT was the most steady gene, which could be the most effective inner referral gene for the expression evaluation of flower advancement in L. discolor. Conclusions: The results of this study will contribute to the molecular biology research of flower development in L. discolor and closely related species.

Functional characterization of Arabidopsis hydroxynitrile lyase in response to abiotic stress and the regulation of flowering time.

BACKGROUND: Hydroxynitrile lyases (HNLs) are a class of hydrolytic enzymes from a wide range of sources, which play crucial roles in the catalysis of the reversible conversion of carbonyl compounds derived from cyanide and free cyanide in cyanogenic plant species. HNLs were also discovered in non-cyanogenic plants, such as Arabidopsis thaliana, and their roles remain unclear even during plant growth and reproduction. METHODS AND RESULTS: The pattern of expression of the HNL in A. thaliana (AtHNL) in different tissues, as well as under abiotic stresses and hormone treatments, was examined by real-time quantitative reverse transcription PCR (qRT-PCR) and an AtHNL promoter-driven histochemical ß-glucuronidase (GUS) assay. AtHNL is highly expressed in flowers and siliques, and the expression of AtHNL was dramatically affected by abiotic stresses and hormone treatments. The overexpression of AtHNL resulted in transgenic A. thaliana seedlings that were more tolerance to mannitol and salinity. Moreover, transgenic lines of A. thaliana that overexpressed this gene were less sensitive to abscisic acid (ABA). Altered expression of ABA/stress responsive genes was also observed in hnl mutant and AtHNL-overexpressing plants, suggesting AtHNL may play functional roles on regulating Arabidopsis resistance to ABA and abiotic stresses by affecting ABA/stress responsive gene expression. In addition, the overexpression of AtHNL resulted in earlier flowering, whereas the AtHNL mutant flowered later than the wild type (WT) plants. The expression of the floral stimulators CONSTANS (CO), SUPPRESSOR OF OVER EXPRESSION OF CO 1 (SOC1) and FLOWERING LOCUS T (FT) was upregulated in plants that overexpressed AtHNL when compared with the WT plants. In contrast, expression of the floral repressor FLOWERING LOCUS C (FLC) was upregulated in AtHNL mutants and downregulated in plants that overexpressed AtHNL compared to the WT plants. CONCLUSION: This study revealed that AtHNL can be induced under abiotic stresses and ABA treatment, and genetic analysis showed that AtHNL could also act as a positive regulator of abiotic stress and ABA tolerance, as well as flowering time.

Isolation and characterisation of sesquiterpene synthase from aromatic orchid Phalaenopsis bellina (Rchb.f.) Christenson.

BACKGROUND: Phalaenopsis bellina, an orchid native to Borneo, is renowned for its unique appearance. It releases distinct fragrances, which have been linked to the presence of terpenoids. However, the identification and study of sesquiterpene synthase in P. bellina remain limited. In this study, we examines the functional characterisation of terpene synthase (TPS) from P. bellina, known as PbTS, through recombinant protein expression and its manifestation in the flower. METHODS AND RESULTS: Gene annotation of PbTS revealed that the inferred peptide sequence of PbTS comprises 1,680 bp nucleotides encoding 559 amino acids with an estimated molecular mass of 65.2 kDa and a pI value of 5.4. A similarity search against GenBank showed that PbTS shares similarities with the previously published partial sequence of P. bellina (ABW98504.1) and Phalaenopsis equestris (XP_020597359.1 and ABW98503.1). Intriguingly, the phylogenetic analysis places the PbTS gene within the TPS-a group. In silico analysis of PbTS demonstrated stable interactions with farnesyl pyrophosphate (FPP), geranyl pyrophosphate (GPP), and geranylgeranyl pyrophosphate (GGPP). To verify this activity, an in vitro enzyme assay was performed on the PbTS recombinant protein, which successfully converted FPP, GPP, and GGPP into acyclic sesquiterpene ß-farnesene, yielding approximately 0.03 mg/L. Expressional analysis revealed that the PbTS transcript was highly expressed in P. bellina, but its level did not correlate with ß-farnesene levels across various flowering time points and stages. CONCLUSION: The insights gained from this study will enhance the understanding of terpenoid production in P. bellina and aid in the discovery of novel fragrance-related genes in other orchid species.