Effective phytoremediation depends on plant species that combine high biomass production with efficient P accumulation and tolerance to elevated nutrient loads. Therefore, this study aimed to compare the P removal efficiency (PRE), uptake capacity, and toxicity threshold among four Azolla samples from three species (A. pinnata, A. caroliniana, and A. filiculoides) grown under in high-P media. Growth responses, P, Fe, N, and Zn content, and PRE were measured. All species showed similar growth and P uptake responses up to 5 mM P. Beyond this concentration, species-specific responses were observed: A. pinnata became toxic and died while A. filiculoides (Utr.) tolerated up to 25 mM. In contrast, A. caroliniana and A. filiculoides (Can) maintained stable growth even under higher P exposure conditions, despite no further increase in tissue P concentration. This pattern suggests a regulated response to P accumulation, rather than dilution or luxury consumption (nutrient uptake and accumulation beyond required levels without further increase in biomass). P removal efficiency decreased significantly at concentrations above 5 mM and remained unchanged at higher P supplies, indicating that the contaminant concentration significantly affects the removal efficiency. Overall, the results indicate significant inter- and intra-specific variation in P toxicity, uptake and removal in Azolla. Therefore, this highlights the importance of selecting the appropriate species and considering contaminant concentration for efficient phytoremediation.

Glycyrrhiza glabra L., belonging to the Fabaceae family, is an important source of pharmacologically active compounds, especially glycyrrhizin. Hairy root culture is an efficient approach for the stable biosynthesis of secondary metabolites. In this study, three experimental sets were performed to optimize hairy root establishment. Initially, the effect of explant type and Agrobacterium rhizogenes strain on the effectiveness of hairy root induction was evaluated. The maximum transformation frequency (86 Not applicable.

Milk thistle (Silybum marianum), a medicinal plant from the Asteraceae family, is widely known for its high-value secondary metabolite, silymarin, a flavonolignan with potent hepatoprotective and antioxidant properties. This study aimed to enhance silymarin production by eliciting cell suspension cultures with methyl jasmonate (MeJA) and salicylic acid (SA). Initially, callus induction was optimized using leaf explants treated with kinetin, 2,4-Dichlorophenoxyacetic acid (2,4-D), and picloram, achieving maximum proliferation with 0.8 mg/L kinetin, 0.1 mg/L 2,4-D, and 2 mg/L picloram. Subsequent elicitation of suspension cultures revealed that 0.05 mM MeJA and 0.25 mM SA resulted in the highest silymarin accumulation, as confirmed by High-performance liquid chromatography (HPLC) analysis. Additionally, quantitative polymerase chain reaction-based gene expression profiling demonstrated that MeJA and SA differentially regulated the transcription of Phenylalanine ammonia-lyase (PAL), Cinnamyl Alcohol Dehydrogenase (CAD), and Chalcone Isomerase (CHI), key genes involved in the phenylpropanoid pathway, with distinct temporal expression patterns. These findings underscore the efficacy of MeJA and SA as potent elicitors for enhancing silymarin biosynthesis, offering a promising biotechnological strategy for optimizing secondary metabolite production in medicinal plants.

Kelussia odoratissima Mozaff (celery) is a valuable endemic spice and aromatic plant of the Apiaceae family, rich in secondary metabolites. This study aimed to optimize in vitro regeneration methods and LEDs to enhance the phenolic compounds and chemical composition of Kelussia. Various combinations and concentrations of plant growth regulators (PGRs) were tested using three types of explants: cotyledons, hypocotyls, and zygotic embryos. In the second part of the study, five light treatments were tested: viz. red, blue, red‒blue, white, and control. Secondary volatile oil compounds were analyzed using solid‒phase microextraction (SPME) arrow to assess changes under each light condition. HPLC analysis identified the principal phenolic and flavonoid compounds in the different explants, such as caffeic acid, chlorogenic acid, and rutin. In particular, the highest concentration of phenolic compounds (caffeic acid: 43.09 µg/g) was detected in zygotic embryo explants treated with 2–4-D at 0.5 mg/L and BAP at 0.25 mg/L. The SPME analysis revealed that (E)-farnesene (32.09

Heavy metal contamination in aquatic ecosystems poses serious environmental and health risks. Azolla caroliniana, a promising candidate for phytoremediation, has the potential to absorb heavy metals like lead (Pb). However, limited information is available on the enzymatic and genetic responses of A. caroliniana under Pb stress. This study investigates the plant's phytoremediation capacity by analyzing antioxidant enzyme activity and gene expression under lead (II) acetate [Pb(C2H3O2)2] concentrations (0, 500, 750, 1,000 µM) over three time points (days 2, 4, and 6). The results showed that with increasing Pb concentration, antioxidant enzyme activity increased. Chlorophyll content increased by 25% at 1,000 µM Pb, whereas carotenoid and anthocyanin levels decreased by 233% and 30%, respectively. Total protein content declined by 90%. Additionally, SOD and CAT activities increased by 28%, while APX activity rose by 25%. Gene expression analysis revealed that genes associated with antioxidant enzymes CAT (94% decrease), APX (64% decrease), SOD (40% decrease), GR (8% increase), and PPO (93% decrease) as well as anthocyanin biosynthesis genes C4H (56% decrease) and CHS (87% decrease) were significantly downregulated at the highest Pb concentration in the later stages, indicating a critical adaptation phase. Observed gene expression fluctuations in the later stages may result from A. caroliniana's dynamic stress response, where initial upregulation of antioxidant defense genes suggests an attempt to mitigate oxidative stress, followed by metabolic adjustments leading to variations in gene expression levels. Lead uptake peaked on day 2 but significantly declined by 42% on day 6, likely due to cellular saturation, activation of detoxification mechanisms, or lead translocation into the growth medium. These findings highlight the potential of A. caroliniana as an effective phytoremediator for Pb-contaminated water bodies.

Greenhouses located at high latitudes and in cloudy areas often experience a low quality and quantity of light, especially during autumn and winter. This low daily light integral (DLI) reduces production rate, quality, and nutritional value of many crops. This study was conducted on Sakhiya RZ F1 tomato plants to evaluate the impact of LED lights on the growth and nutritional value of tomatoes in a greenhouse with low daily light due to cloudy weather. The treatments included LED growth lights in three modes: top lighting, intra-canopy lighting, and combined top and intra-canopy lighting. The results showed that although the combined top and intra-canopy lighting reached the maximum increase in tomato yield, exposure to intra-canopy LED lighting alone outperformed in tomato fruit yield increase (28.46%) than exposure to top LED lighting alone (12.12%) when compared to no supplemental lighting during the entire production year. Intra-canopy exposure demonstrated the highest increase in tomato lycopene (31.3%), while top and intra-canopy lighting exhibited the highest increase in vitamin C content (123.4%) compared to the control. The LED light treatment also had a very positive effect on the expression of genes responsible for metabolic cycles, including Psy1, LCY-β, and VTC2 genes, which had collinearity with the increase in tomato fruit production.

Cadmium (Cd) contamination poses significant risks to agricultural productivity and human health, particularly through its accumulation in staple crops such as bread wheat (Triticum aestivum L.). This study evaluated Cd accumulation and tolerance among six bread wheat cultivars exposed to six Cd concentrations (0, 2.5, 5, 10, 15, 20, and 25 mg kg-1 soil). Phenotypic assessments and quantitative real-time PCR (qRT-PCR) were conducted to analyze the expression patterns of TaNRAMP and TaZIP genes in various tissues and developmental stages of wheat, which play crucial roles in Cd uptake and transport. Results demonstrated significant variability in Cd accumulation. The Barat cultivar exhibited the lowest accumulation in grain (ranging from 0.21 to 8.8 mg kg-1) and the highest tolerance. In contrast, Kavir and Pishtaz displayed elevated Cd levels in both grain and straw, while Parsi accumulated more Cd in straw at lower concentrations (56.9 mg kg-1 in Cd concentration of 10 mg kg-1 soil). The gene expression analysis revealed that most cultivars showed increased expression of TaNRAMP genes, particularly TaNRAMP2 in Cd concentration of 10 mg kg-1 soil, which facilitates Cd uptake from the soil, and TaZIP genes, such as TaZIP4 and TaZIP7, involved in transporting Cd within the plant. Notably, the expression of TaZIP1 was significantly lower in cultivars with high Cd accumulation, suggesting a potential regulatory mechanism for Cd tolerance. Furthermore, cultivars exhibiting higher Cd levels correlated with increased expression of stress-responsive genes, indicating a broader response to Cd stress. These findings highlight Barat's potential for bread-making applications due to its low Cd accumulation, while Morvarid and Pishtaz which show reduced Cd content in the straw even under high Cd exposure are better suited for animal feed. This research underscores the genetic variability of wheat cultivars in response to Cd stress and provides essential insights into the molecular mechanisms underlying Cd accumulation, offering valuable information for breeding programs aimed at developing Cd-tolerant varieties to ensure food security in contaminated regions.

Glycyrrhizin is a valuable secondary metabolite with high medicinal properties, which is known as the most important compound in the licorice (Glycyrrhiza glabra L.). Considering the problems in growing plants in the field, the use of tissue culture techniques, especially hairy root culture, makes the production of glycyrrhizin simpler and more affordable. The influence of Agrobacterium rhizogenes strains on the efficiency of hairy root induction showed that the highest hairy roots were observed using MSU strain (80%). Also, the effect of explant type (leaf, stem and petiole) and age (two-week-old, three-week-old and four-week-old), light conditions (24 hours of darkness and 16 hours of light-8 hours of darkness) and carbohydrate source (3% glucose and 3% sucrose) showed that the production of hairy roots from this medicinal plant depends on the type and age of explant, light treatment and the composition of the culture medium and the highest hairy roots induction was observed in leaf explant (80%), three-week-old leaf (76.66%) and MS medium containing 3% sucrose (86.66%) and treatment of 24 hours of darkness (86.66%). The amount of glycyrrhizin in hairy root lines treated by 100 µM methyl jasmonate (MJA) after 24 hours increased in the no cutting treated line and decreased in the cutting treated lines compared to the control line. In conclusion, elicitation of G. glabra hairy root culture is a suitable platform for achieving high glycyrrhizin yields.

Improving flower yield through lengthening flowering duration is a primary breeding objective in saffron (Crocus sativus L.). Asexual reproduction in saffron limits biodiversity and conventional breeding. Hence, eliciting flowering-related gene expression by plant growth regulators is one way to achieve this aim. The phytohormones methyl jasmonate (MeJA) and 6-benzyl amino purine (BAP) signals are received by the MADs-box gene family. In this study, to elucidate the role of phytohormones on flower development, plant were treated with BAP (0 and 5 mg L−1), and methyl jasmonate (MeJA) (0, 20, and 100 mM) at three developmental stages of the saffron life cycle. Then, the expression of the SHORT VEGETATIVE PHASE (CsSVP) gene as a MADS-box gene family was assessed in the saffron corm. The activities of antioxidant enzymes, soluble sugar, starch content, and soluble protein content were also measured in corm, leaf, and root tissues. The application of MeJA and BAP treatments resulted in down-regulation of CsSVP expression in the corm during dormancy. At the dormancy stage, catalase, peroxidase activity decreased, and ascorbate peroxidase activity increased following MeJA treatment. In contrast, an increment in catalase and peroxidase activity and reduction of ascorbate peroxidase activity were observed after treatment with MeJA during the flowering stage. This change in enzyme activity is most likely due to flowering, which demands the re-allocation of resources. As flowering is a process heavily influenced by the environment, plants treated with MeJA, which may mimic environmental stress, showed changes in antioxidant enzyme activity. Overall, these results suggested that MeJA and BAP treatments play a significant role in the vegetative-to-reproductive phase change in saffron.

Almond is a stone fruit crop belonging to the Rosaceae family, cultivated in the temperate region of the world for its high nutritive valued fruits. In today’s marketplace, the grower profit is directly related to input traits such as large fruit size. To elucidate the genetic control of seed size in almonds, we use the xenia phenomenon. For this purpose, in this study, almond cultivars ‘Sefid’ and ‘Mamaee Pooya’ were pollinated with pollen from almond cultivars ‘Mamaee’ (large seeds) and a genotype ‘Orientalis’ (small seeds). The seed quantitative traits showed pollination with ‘Mamaee’ increased these traits of each female parent compared to pollination with ‘Orientalis’. Comparative profiling demonstrated 258 differentially expressed genes (DEGs) between the two assayed RNA-Seq samples (one sample containing a pool of seeds of ‘Sefid’ × ‘Mamaee’ and the other a pool of seeds of ‘Sefid’ × ‘Orientalis’). Which among them 150 were up-regulated and 108 were down-regulated. Gene ontology (GO) annotation analysis revealed that DEGs were associated with hormone signaling, metabolite synthesis and cell communication. KEGG pathway analysis on 256 DEGs showed that many genes are engaged in the control of kernel size. Also, analysis of protein–protein interaction network displayed that several genes such as PduBEN1 (Protein BRI1-5 ENHANCED 1), PduSUS2 (Sucrose synthase 2), PduGA2OX2 (Gibberellin 2-beta-dioxygenase 2), PduABCG25 (ABC transporter G family member 25), PduGASA1 (gibberellic acid-stimulated Arabidopsis) and PduCYP72C1 (cytochrome p450 72c1) had a remarkable impact on kernel size. This study has provided considerable insights into the complicated regulatory mechanism underlying the kernel size trait in almonds.

This study aimed to evaluate the effect of crop load on the fruit quality traits especially nutritional and nutraceutical properties of two cultivars of greenhouse tomato (Solanum lycopersicum L.) i.e. ‘Grandella’ and ‘Isabella’. The crop load treatments were: saving one fruit per truss (1F), two fruits per truss (2F), three fruits per truss (3F), and no fruit thinning (control). The results showed that decreasing crop load increased the dry matter content, glucose, fructose, and sucrose concentrations of fruits because of less competition between fruits of each truss and higher relative expression level of tomato hexose transporter genes (LeHTs). Higher transport of sugars toward fruits of 1F resulted in high secondary metabolites concentration. The high secondary metabolites concentration with high fruit load in control that has low soluble sugar availability may result from the opportunity to take advantage of the excess pool of carbon to build cost-free carbon-based secondary metabolites. A reduced crop load would be a recommended management strategy to improve greenhouse tomato quality.

Many studies have investigated the role of miRNAs on the yield of various plants, but so far, no report is available on the identification and role of miRNAs in fruit and seed development of almonds. In this study, preliminary analysis by high-throughput sequencing of short RNAs of kernels from the crosses between almond cultivars 'Sefid' × 'Mamaee' (with small and large kernels, respectively) and 'Sefid' × 'P. orientalis' (with small kernels) showed that the expressions of several miRNAs such as Pdu-miR395a-3p, Pdu-miR8123-5p, Pdu-miR482f, Pdu-miR6285, and Pdu-miR396a were significantly different. These miRNAs targeted genes encoding different proteins such as NYFB-3, SPX1, PGSIP3 (GUX2), GH3.9, and BEN1. The result of RT-qPCR revealed that the expression of these genes showed significant differences between the crosses and developmental stages of the seeds, suggesting that these genes might be involved in controlling kernel size because the presence of these miRNAs had a negative effect on their target genes. Pollen source can influence kernel size by affecting hormonal signaling and metabolic pathways through related miRNAs, a phenomenon known as xenia.

Water ferns of the genus Azolla and the filamentous cyanobacteria Nostoc azollae constitute a model symbiosis that enabled the colonization of the water surface with traits highly desirable for the development of more sustainable crops: their floating mats capture CO2 and fix N2 at high rates using light energy. Their mode of sexual reproduction is heterosporous. The regulation of the transition from the vegetative phase to the spore forming phase in ferns is largely unknown, yet a prerequisite for Azolla domestication, and of particular interest as ferns represent the sister lineage of seed plants. Sporocarps induced with far red light could be crossed so as to verify species attribution of strains from the Netherlands but not of the strain from the Anzali lagoon in Iran; the latter strain was assigned to a novel species cluster from South America. Red-dominated light suppresses the formation of dissemination stages in both gametophyte- and sporophyte-dominated lineages of plants, the response likely is a convergent ecological strategy to open fields. FR-responsive transcripts included those from MIKCC homologues of CMADS1 and miR319-controlled GAMYB transcription factors in the fern, transporters in N. azollae, and ycf2 in chloroplasts. Loci of conserved microRNA (miRNA) in the fern lineage included miR172, yet FR only induced miR529 and miR535, and reduced miR319 and miR159. Phylogenomic analyses of MIKCC TFs suggested that the control of flowering and flower organ specification may have originated from the diploid to haploid phase transition in the homosporous common ancestor of ferns and seed plants.

Saffron is the world highest-priced spice because its production requires intensive hand labour. Reduce saffron production costs require containerised plant production under controlled conditions and expand the flowering period. Controlling the flowering process and identify the factors involved in saffron flowering is crucial to introduce technical improvements. The research carried out so far in saffron has allowed an extensive knowledge of the influence of temperature on the flower induction, but the molecular mechanisms controlling flowering induction processes are largely unknown. The present study is the first conducted to isolate and characterize a regulator gene of saffron floral induction the Short Vegetative Phase (SVP) gene, which represses the floral initiation genes in the temperature response pathway, which involved in saffron flower induction. The results obtained from both phylogenetic analysis and T-coffee alignment confirms that the isolated sequence belongs to the SVP gene clades of MADS-box gene family. Gene expression analysis in different developmental stages revealed the highest expression of SVP transcript (CsSVP) during the dormancy and the vegetative stages, but decrease when flower development initiated and it was the least in late September when flower primordia are developed. Furthermore, its expression increased in the apical bud when corms are storage at 9–10 ºC, thus inhibiting flower induction. Additionally, comparison of the CsSVP transcript in apical buds from big and small corms, differing in their flowering capacity, indicates that the CsSVP transcript is present only in vegetative buds. Taken together, these results suggested inhibitory role of the SVP gene.

ABSTRACT Azolla ferns and the filamentous cyanobacteria Nostoc azollae constitute a model symbiosis that enabled colonization of the water surface with traits highly desirable for development of more sustainable crops: their floating mats capture CO 2 and fixate N 2 at high rates phototrophically. Their mode of sexual reproduction is heterosporous. Regulation of the transition from vegetative to spore-forming phases in ferns is largely unknown, yet a pre-requisite for Azolla domestication, and of particular interest since ferns represent the sister lineage of seed plants. Far-red light (FR) induced sporocarp formation in A. filiculoides . Sporocarps obtained, when crossed, verified species attribution of Netherlands strains but not Iran’s Anzali lagoon. FR-responsive transcripts included CMADS1 MIKC C -homologues and miRNA-controlled GAMYB transcription factors in the fern, transporters in N . azollae , and ycf2 in chloroplasts. Loci of conserved miRNA in the fern lineage included miR172, yet FR only induced miR529 and miR535, and reduced miR319 and miR159. Suppression of sexual reproduction in both gametophyte and sporophyte-dominated plant lineages by red light is likely a convergent ecological strategy in open fields as the active control networks in the different lineages differ. MIKC C transcription factor control of flowering and flower organ specification, however, likely originated from the diploid to haploid phase transition in the homosporous common ancestor of ferns and seed plants.

This study investigated the effect of drought stress on the amount of phenolic and flavonoid compounds as well as H2O2 and malondialdehyde (MDA) in Achillea pachycephala. The expression patterns of the key genes and their molecular mechanisms in the phenylpropanoid pathway (PAL, CHS, CHI, F3H, F3'H, F3'S'H, FLS) were also assessed during drought stress using quantitative real-time polymerase chain reaction (qRT-PCR). The samples were harvested at 0, 7, 14, 21 and 28 days after exposure to drought stress. High-performance liquid chromatography (HPLC) analysis was performed to determine the changes of phenolic and flavonoid compounds - chlorogenic acid, caffeic acid, rutin, luteolin-7-O-glycoside, 1,3-dicaffeoylquinic acid, apigenin-7-O-glycoside, luteolin, apigenin and kaempferol - during stress conditions. Concentrations of most of the compounds increased with increasing drought stress duration. Most of the phenolic acids continued to accumulate with increasing duration of stress, while flavonoids dramatically decreased at day 28 of stress. Chlorogenic acid was the most abundant phenolic acid (4.97 mg/100 g dry weight [DW]) at the beginning of the experiment, while it decreased at day 7 and increased again at day 21. However, different trends were observed for some flavonoids, such as luteolin and apigenin. At the beginning of stress treatment, high accumulation of free radicals (H2O2) and lipid peroxidation (MDA) led to elevated expression of most of the flavonoid genes. MDA increased from 22.66 to 43.28 mu mol g(-1) DW at day 28. CHS gene expression was elevated at day 7, while chi gene expression remained unchanged. At the end of the stress period, most of the flavonoid concentrations and expression of the relevant genes also increased. The results can facilitate selection of appropriate drought conditions to obtain the highest levels of flavonoids such as luteolin and apigenin and phenolic compounds such as chlorogenic acid for improved health benefits.

Azolla is a floating aquatic fern, having amazing capacity for concentrating toxic heavy metals. Metallothioneins (MTs) and phytochelatins (PCs) are well-defined heavy metal-binding ligands in plants. Bioaccumulation potential of different Azolla species varies according to their heavy metal ions. Therefore, the accumulation of Ni, Zn, Cu, and Cd was studied in A. pinnata, A. filiculoides, and a sample taken from Anzali wetland. Moreover, the expression of metallothionein and phytochelatin synthase encoding genes was examined at different metal concentrations. The highest level of Cu and Cd absorption was detected in A. pinnata, while the maximum amount of Ni and Zn absorption was observed in A. filiculoides and the sample taken from Anzali, respectively. The MT2 and PCS1 gene expression patterns were significantly induced by the heavy metal treatments, confirming their roles in phytoremediation potential of Azolla. However, as the results concerning heavy metal accumulation and gene expression vary in different species, only specific species of Azolla can be used for special purposes. It can be concluded that the Azolla is a good candidate for phytoremediation purposes, and the formation of phytochelatin-heavy metal complexes and their sequestration in vacuole are the main processes influencing susceptibility of Azolla to heavy metals.

In order to examine the relationship between proline and cold stress in Iranian (local) genotype of petunia, p5cs gene was transferred to petunia through Agrobacterium-mediated transformation. The leaf discs from four weeks and shoot apices from 7-day-old petunia in vitro plants were co-cultivated with Agrobacterium strain LBA4404 harboring a plasmid pBI121 as the vector system for transformation of petunia. pBI121 plasmid containing β-glucuronidase (gus) gene as a reporter gene and Δ1-pyrroline-5-carboxylate synthetase (p5cs) gene and neomycin phosphotransferase (nptII) gene were used as a selectable marker. The co-cultivated leaf discs and shoot apices were transferred to the selective medium and thereafter to proliferation medium respectively. To confirm transformation, regenerated plants were subjected to the polymerase chain reaction (PCR), GUS histochemical and proline assays. The results confirmed the presence of the gus and p5cs gene in the genome of all transformants and the transformed plants were more tolerant (674.87 μg/g fresh leaves) than the wild types under stress conditions.Plant Tissue Cult. & Biotech. 28(1): 35-44, 2018 (June)