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• 2,4-D pulse at 20 µM and 30 µM induces direct somatic embryogenesis on the embryonic axis of Plinia peruviana. • 26.7 % of the induction rate of direct somatic embryogenesis was obtained using 20 µM of 2,4-D for 5 days. • Germination of embryonic axes is inversely proportional to the auxin concentration in Plinia peruviana. • Additional improvements to the protocol are still needed to improve embryo conversion into plantlets. Plinia peruviana (Myrtaceae) is a fruit tree species endemic to Brazil with high importance to the food and pharmacological industries. Despite the need for vegetative propagation of selected genotypes for genetic breeding or fruit production, efficient propagation methods for the species have not yet been determined. This study aimed to advance the establishment of a protocol for inducing somatic embryogenesis from mature P. peruviana seeds. Three concentrations of 2,4-dichlorophenoxyacetic acid (10, 20, and 30 µM) in four auxinic pulses (1, 3, 5, and 7 days of exposure times) were tested. 20 µM and 30 µM of 2,4-D promoted the highest callogenesis (20–40 %), regardless of the exposure period, but calli did not present embryogenic potential. The formation of somatic embryos occurred asynchronously directly on the embryonic axes and was dependent on adding 2,4-D to the culture medium. The highest induction rate of direct somatic embryogenesis (26.7 %) was obtained using 20 µM of 2,4-D for 5 days of exposure. The light and electronic microscopic analyses allowed the characterization of the embryos at different stages of development and identified some abnormalities. Additional improvements to the protocol are still needed, including testing different culture media since no conversion and formation of complete plantlets was achieved. [ABSTRACT FROM AUTHOR]

Cucumber (Cucumis sativus L.) is a genetically diverse group of vegetables with various cultivars having distinct traits. This study optimized somatic embryogenesis in anther-derived calli of selected commercial cucumber cultivars (Local Khera, Champion, and CP 001). Therefore, the experiment investigated the impact of 2,4-D and benzyl 6-aminopurine (BAP) treatments on embryo formation. The anthers collected from different-sized floral buds sustained culturing in various concentrations of 2,4-D and BAP (0, 0.5, 1, 1.5, 2, 3, and 4 mg L-1). It was evident that calli induction in cucumber cultivars received significant stimuli from 2,4-D and BAP concentrations and dark culture conditions during calli culture. The maximum calli induction (51%) was prominent in anthers of cv. CP 001 at a higher level of 2,4-D in small-sized floral buds. However, the anthers of cucumber cultivar Local Khera (59.72%) performed better for calli induction than Champion (57.14%) and CP 001 (51.43%). The highest embryogenesis appeared in anther-derived calli of cultivar Local Khera (12%) under light culture conditions in tiny flower buds. Meanwhile, maximum (8%) embryo formation observed at a higher level of 2,4-D (4 mg L-1) resulted in cultivars Champion and CP 001 under dark conditions. In conclusion, from the tested treatments, applying the highest level of 2,4-D and BAP at 4 mg L-1 was more effective than other treatments, including the control. However, more calli induction was noteworthy under dark culture conditions, and maximum embryo formation occurred under light culture conditions. [ABSTRACT FROM AUTHOR]

The green synthesis of AgNPs from the calli extract of Centella asiatica was confirmed by the detailed analysis of UV–Vis spectroscopy, Scanning electron microscopy (SEM), Energy dispersive X-ray analysis (EDAX), Transmission electron microscopy (TEM), Fourier Transform Infrared Spectroscopy (FT-IR) which revealed the size, shape, chemical composition and functional groups involved in the AgNPs formation. The in-vitro raised cowpea plants were given root dip treatment in Murashige and Skoog (MS) suspension medium supplemented with LC 50 - 75 mg/mL AgNPs followed by their transfer to glass house conditions for acclimatization. To evaluate the efficiency of root dip treatment, the acclimatized plants were inoculated with 2000 J2s of M. arenaria and the plants were observed on weekly basis. A differential growth response was observed from the cowpea plants under glass house conditions. A treatment plan designed showed that P4 plants showed a significant defense response against the biotic stress of M. arenaria as indicated by the percentage response of several growth parameters such as- plant height of 49.22 ± 0.89 %, plant fresh mass of 46.12 ± 0.83 %, plant dry mass of 9.3 ± 0.17 % and nodule number of 38 ± 0.68 % were reported in P4 plants after 6 weeks of J2s inoculation. Confocal microscopy of root sections of P3 cowpea plants revealed the presence of M. arenaria J2s in the cortical and provascular regions, however, in case of P4 plants an intact cellular structure was maintained. The application of AgNP didn't show any toxic effect on the growth of plant. [Display omitted] • The UV peaks at 430 nm and FTIR analysis displayed several prominent functional groups. • Toxicity of green synthesized AgNPs showed a concentration dependent nematicidal effect. • Biogenic nano-silvers exhibited significant growth enhancement in cowpea plants. • Biogenic nano-silvers exhibited significant defense response against J2s of M. arenaria. [ABSTRACT FROM AUTHOR]

Crocus scepusiensis (Rehm. & Woł.) Borbás ex Kulcz., a critically endangered herbaceous plant which serves as a valuable source of bioactive compounds found across Europe and Asia. The aim of this study was to produce a calli from two different plant parts (leaf and shoot tip) for the critically endangered C. scepusiensis through tissue culture techniques, characterize the resulting calli through chemical profiling, with a focus on identifying key phytoconstituents, and lay the groundwork for future research on the biological activities of these calli extracts. Leaf disc and micro shoot tip explants were cultured on Murashige and Skoog (MS) medium supplemented with various concentrations of 6-benzylaminopurine (BA) and α-naphthaleneacetic acid (NAA) to induce organogenic calli. The resulting calli exhibited distinct biochemical profiles. Moreover, a phytochemical analysis was conducted to compare the metabolite composition of callus 1 (derived from leaf discs) and callus 2 (derived from shoot tips). Callus 1 displayed a higher total phenolic content (30.3558 ± 1.3564 mg (GAE)/g) compared to callus 2 (29.1543 ± 0.9754 mg (GAE)/g). Similarly, callus 1 exhibited a greater total flavonoid content (26.0089 ± 1.8029 mg (RE)/g) than callus 2 (18.4464 ± 1.4797 mg (RE)/g). Liquid chromatography-photodiode array-electrospray ionization-tandem mass spectrometry (LC-PDA-ESI-MS/MS) analysis revealed the presence of 26 and 25 constituents in callus 1 and 2, respectively. Fourteen and thirteen of these identified compounds have been previously reported in other Crocus species, with 22 constituents common to both calli. Twelve constituents were reported here in Crocus for the first time as far as we know.Key message: Tissue culture successfully produced Crocus scepusiensis calli with distinct phytoconstituents profiles, potentially offering novel sources of bioactive compounds. [ABSTRACT FROM AUTHOR]

This study aimed to optimize embryogenic callus formation and transformation of the CP57-614 sugarcane variety using particle bombardment. Young leaf explants were cultured on modified Murashige and Skoog (MS) medium containing different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D; auxin) (1, 2, and 4 mg L-1) and two Dicamba concentrations (4 and 6 mg L-1). The highest callus induction rate was achieved with 2 mg L-1 of 2,4-D. Subsequently, particle bombardment was used to introduce genetic constructs containing the uidA reporter gene driven by three promoters (CaMV35S, ubiquitin, and actin1) into the embryogenic calli. We assessed the impact of various biological, physical, and DNA parameters, including bombardment pressure, target distance, particle size, DNA amount, and osmotic pretreatment, through histochemical GUS assay to quantify blue spots. Results showed that the most effective combination involved bombardment at 1100 psi, 6 cm target distance, 1 μm gold particles coated with 1 μg DNA, under a vacuum of 25 inHg. Additionally, the best osmotic pretreatment utilized a solution containing 0.2 M sorbitol and 0.2 M mannitol. Notably, the construct driven by the ubiquitin promoter resulted in the highest transient expression of the uidA gene, suggesting its superiority for gene expression in this system. [ABSTRACT FROM AUTHOR]

Abstract The study aimed to edit ethylene (ET) biosynthesis genes [1-aminocyclopropane-1-carboxylic acid (ACC) synthetase 1 (ACS1) and ACC oxidase 1 (ACO1)] in carnation using the CRISPR/Cas9 ribonucleoprotein (RNP) complex system. Initially, the conserved regions of the target genes (ACS1 and ACO1) were validated for the generation of different single guide RNAs (sgRNAs), followed by the use of an in vitro cleavage assay to confirm the ability of the sgRNAs to cleave the target genes specifically. The in vitro cleavage assay revealed that the sgRNAs were highly effective in cleaving their respective target regions. The complex of sgRNA: Cas9 was directly delivered into the carnation protoplast, and the target genes in the protoplast were deep-sequenced. The results revealed that the sgRNAs were applicable for editing the ET biosynthesis genes, as the mutation frequency ranged from 8.8 to 10.8% for ACO1 and 0.2–58.5% for ACS1. When sequencing the target genes in the callus derived from the protoplasts transformed with sgRNA: Cas9, different indel patterns (+ 1, − 1, and − 8 bp) in ACO1 and (− 1, + 1, and + 11) in ACS1 were identified. This study highlighted the potential application of CRISPR/Cas9 RNP complex system in facilitating precise gene editing for ET biosynthesis in carnation.

BackgroundAn efficient in vivo transient transfection system using protoplasts is an important tool to study gene expression, metabolic pathways, and multiple mutagenesis parameters in plants. Although rice protoplasts can be isolated from germinated seedlings or cell suspension culture, preparation of those donor tissues can be inefficient, time-consuming, and laborious. Additionally, the lengthy process of protoplast isolation and transfection needs to be completed in a single day.ResultsHere we report a protocol for the isolation of protoplasts directly from rice calli, without using seedlings or suspension culture. The method is developed to employ discretionary pause points during protoplast isolation and before transfection. Protoplasts maintained within a sucrose cushion partway through isolation, for completion on a subsequent day, per the first pause point, are referred to as S protoplasts. Fully isolated protoplasts maintained in MMG solution for transfection on a subsequent day, per the second pause point, are referred to as M protoplasts. Both S and M protoplasts, 1 day after initiation of protoplast isolation, had minimal loss of viability and transfection efficiency compared to protoplasts 0 days after isolation. S protoplast viability decreases at a lower rate over time than that of M protoplasts and can be used with added flexibility for transient transfection assays and time-course experiments. The protoplasts produced by this method are competent for transfection of both plasmids and ribonucleoproteins (RNPs). Cas9 RNPs were used to demonstrate the utility of these protoplasts to assay genome editing in vivo.ConclusionThe current study describes a highly effective and accessible method to isolate protoplasts from callus tissue induced from rice seeds. This method utilizes donor materials that are resource-efficient and easy to propagate, permits convenience via pause points, and allows for flexible transfection days after protoplast isolation. It provides an advantageous and useful platform for a variety of in vivo transient transfection studies in rice.

In order to study the chemical compounds of the calli induced from Maytenus hookeri, the dried and fresh cells of the calli were extracted by heat refluxing and cold soaking method, respectively. The chemical constituents were purified through various column chromatography including silica gel, RP-18 and Sephadex LH-20 from the ethyl acetate extract. According to the NMR and MS analysis, sixteen compounds were identified as wilforlide A ( 1), wilforlide B (2), 3β,22α-dihydroxyolean- 12-en-29-oic acid (3), 22α-hydroxy-3-oxoolean-12-en-29-oic acid (4), 3-epikatonic acid (5), oleana-9(11),12-dien-3-β-ol (6), 22α-hydroxy-3-oxo-12-ursen-30-oic acid (7), triptotriterpenic acid C (8), 2α,3β-dihydroxy-29-friedelanoic acid (9), maytenonic acid (10), 3β-hydroxystigmast-5-en-7-one (11), 7β-hydroxysitosterol (12), stigmast-4-ene-3-one (13), β-sitosterol (14), 1-β-Dglucpotyransyl- 2,6-dimetoxi-4-propenyl-benzene (15) and adenosine (16). All compounds were reported in the calli of M. hookeri and the plant for the first time except 10 and 14. [ABSTRACT FROM AUTHOR]

Abstract Background Prunella vulgaris is medicinally important plant containing high-valued chemical metabolites like Prunellin which belong to family Lamiaceae and it is also known as self-heal. In this research, calli culture were exposed to differential ratios of gold (Au) and silver (Ag) nanoparticles (1:1, 1:2, 1:3, 2:1 and 3:1) along with naphthalene acetic acid (2.0 mg NAA) to investigate its antimicrobial potential. A well diffusion method was used for antimicrobial properties. Results Here, two concentrations (1 and 2 mg/6 µl) of all treated calli cultures and wild plants were used against Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Bacillus atrophaeus, Bacillus subtilis, Agrobacterium tumefaciens, Erwinia caratovora and Candida albicans. Dimethyl sulfoxide (DMSO) and antibiotics were used as negative and positive controls. Here, the calli exposed to gold (Au) nanoparticles (NPs) and 2.0 mg naphthalene acetic acid (NAA) displayed the highest activity (25.7 mm) against Salmonella typhi than other extracts, which was considered the most susceptible species, while Agrobacterium tumefaciens and Candida albicans was the most resistance species. A possible mechanism of calli induced nanoparticles was also investigated for cytoplasmic leakage. Conclusion From the above data it is concluded that Prunella vulgaris is medicinally important plant for the development of anti-microbial drugs using nanotechnology and applicable in various pharmaceutical research.

To increase the production of decorated anthocyanins in potato cell cultures, we knocked out a novel potato gene, named Inducer Silencing of Anthocyanins in Cell culture (StISAC), using CRISPR‐Cas9 editing. Our results provided evidence that mutant cell lines doubled the accumulation level of anthocyanins biosynthesized. Moreover, the production of these important pigments was stabilized over time. Our study overcame important challenges in the efficient biotechnological production of these valuable pigments and reported the function of a novel anthocyanin biosynthesis repressor gene. [ABSTRACT FROM AUTHOR]

Abstract To increase the production of decorated anthocyanins in potato cell cultures, we knocked out a novel potato gene, named Inducer Silencing of Anthocyanins in Cell culture (StISAC), using CRISPR‐Cas9 editing. Our results provided evidence that mutant cell lines doubled the accumulation level of anthocyanins biosynthesized. Moreover, the production of these important pigments was stabilized over time. Our study overcame important challenges in the efficient biotechnological production of these valuable pigments and reported the function of a novel anthocyanin biosynthesis repressor gene.

The method of biological ballistics (biolistic transformation, genetic bombardment) of plants is one of the most modern methods used for direct gene transfer into plant cells. The main advantages of this method include the ability to simultaneously incorporate several target genes into the plant genome, carry out transfer without unnecessary agrobacterial parts and plasmid DNA sequences, and the short time needed to produce transgenic cells. For different plant objects, the efficiency of obtaining transgenic plants by the ballistic method varies from 1 to 3 %. For potato plants, the transformation efficiency is quite low at the moment and the selection of optimal conditions for biolistics is one of the pressing issues of practical biotechnology. This article presents a successful experience of introducing two genes of interest into two potato varieties using the biolistic approach. The results of biolistic transformation experiments are presented for two types of explants: potato internodes and calli of the varieties Aksor and Nevskiy. Of the 862 explants used for transformation, 56 regenerated plants were obtained. PCR screening of transformants revealed one plant with the insertion of the chitinase gene, one with the insertion of the endo-β-1,3-glucanase gene, and co-transformation by both genes was confirmed in four regenerants. The average transformation efficiency for potato explants was 0.7 %. A high number of regenerants (56) as opposed to a low number of transformants (6) reflects an attempt to increase the number of regenerants by using a lower concentration of the selective agent (kanamycin). Although this approach requires more effort, it can be used to produce potato lines with integrated genes of interest for further use in crop breeding. The lines of potato obtained in the current study by introducing two genes associated with the plant response to fungal pathogens will be further assessed for their resistance to fungal diseases and, if successful, will be used in potato crop breeding.

Background: Prunella vulgaris is medicinally important plant containing high-valued chemical metabolites like Prunellin which belong to family Lamiaceae and it is also known as self-heal. In this research, calli culture were exposed to differential ratios of gold (Au) and silver (Ag) nanoparticles (1:1, 1:2, 1:3, 2:1 and 3:1) along with naphthalene acetic acid (2.0 mg NAA) to investigate its antimicrobial potential. A well diffusion method was used for antimicrobial properties. Results: Here, two concentrations (1 and 2 mg/6 µl) of all treated calli cultures and wild plants were used against Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Bacillus atrophaeus, Bacillus subtilis, Agrobacterium tumefaciens, Erwinia caratovora and Candida albicans. Dimethyl sulfoxide (DMSO) and antibiotics were used as negative and positive controls. Here, the calli exposed to gold (Au) nanoparticles (NPs) and 2.0 mg naphthalene acetic acid (NAA) displayed the highest activity (25.7 mm) against Salmonella typhi than other extracts, which was considered the most susceptible species, while Agrobacterium tumefaciens and Candida albicans was the most resistance species. A possible mechanism of calli induced nanoparticles was also investigated for cytoplasmic leakage. Conclusion: From the above data it is concluded that Prunella vulgaris is medicinally important plant for the development of anti-microbial drugs using nanotechnology and applicable in various pharmaceutical research. [ABSTRACT FROM AUTHOR]

Abstract Background An efficient in vivo transient transfection system using protoplasts is an important tool to study gene expression, metabolic pathways, and multiple mutagenesis parameters in plants. Although rice protoplasts can be isolated from germinated seedlings or cell suspension culture, preparation of those donor tissues can be inefficient, time-consuming, and laborious. Additionally, the lengthy process of protoplast isolation and transfection needs to be completed in a single day. Results Here we report a protocol for the isolation of protoplasts directly from rice calli, without using seedlings or suspension culture. The method is developed to employ discretionary pause points during protoplast isolation and before transfection. Protoplasts maintained within a sucrose cushion partway through isolation, for completion on a subsequent day, per the first pause point, are referred to as S protoplasts. Fully isolated protoplasts maintained in MMG solution for transfection on a subsequent day, per the second pause point, are referred to as M protoplasts. Both S and M protoplasts, 1 day after initiation of protoplast isolation, had minimal loss of viability and transfection efficiency compared to protoplasts 0 days after isolation. S protoplast viability decreases at a lower rate over time than that of M protoplasts and can be used with added flexibility for transient transfection assays and time-course experiments. The protoplasts produced by this method are competent for transfection of both plasmids and ribonucleoproteins (RNPs). Cas9 RNPs were used to demonstrate the utility of these protoplasts to assay genome editing in vivo. Conclusion The current study describes a highly effective and accessible method to isolate protoplasts from callus tissue induced from rice seeds. This method utilizes donor materials that are resource-efficient and easy to propagate, permits convenience via pause points, and allows for flexible transfection days after protoplast isolation. It provides an advantageous and useful platform for a variety of in vivo transient transfection studies in rice.

Drought and salinity can be serious problems for agricultural productivity in certain planet areas. Leptosphaeria maculans is the causative agent of the blackleg in crucifer plants. In this work, a novel methodology for studying the effects of fungal metabolites (volatile and non-volatile) on plant calli in the presence of abiotic stresses is presented, by using L. maculans, tomato calli, and drought and salinity stresses. In this way, this study has reported how, under salinity and drought stresses, the growth and vitality of tomato calli is inhibited, increasing its tissues-oxidation and accumulation of ROS. By applying metabolites from L. maculans, the growth of calli treated with non-volatile metabolites showed and increment under salinity and drought conditions. On the other hand, calli treated with volatile metabolites showed an increment in tissues-vitality under salinity and drought conditions. A series of gene expression analysis was also conducted in order to determine the performance of related genes. Results of this study showed that growth related gene expression was induced, together with abiotic stress tolerance gene in response to abscisic acid, AREB1. In addition, the application of volatile metabolites from L. maculans on tomato calli without abiotic stresses increases its growth and vitality, and reduces its oxidation and accumulation of ROS, in accordance with the results of gene expression obtained. The ability of L. maculans metabolites to increase plant tolerance to abiotic stresses could be related to their ability to produce volatile and non-volatile-metabolites, which induce the antioxidant enzyme activity or accumulation of antioxidant compounds, or their ability to increase the expression of ABA-dependent response genes to abiotic stresses.

Rheum ribes L. is a perennial wild species. Young shoots and flower bunches are freshly consumed, and root and rhizomes are generally used for medicinal purposes. The aim of the present study was to improve the callus proliferation protocol for R. ribes L. under in vitro conditions. For callus induction, hypocotyl explants taken from 14-day old plantlets germinated in Murashige and Skoog (MS) media were cultured in MS media with 9 plant growth regulator (PGR) combinations containing 6-benzylaminopurine (BAP) (2, 3, and 4 mg/L) + naphthylacetic acid (NAA) (0.1, 0.5, and 1 mg/L). Then, for callus proliferation, 4 PGR combinations containing NAA (0.2 mg/L) + thidiazuron (TDZ) (0.5, 1, 2, and 3 mg) were used in the first set of experiments, and 36 PGR combinations containing BAP (1, 2, 3, and 4 mg/L) + indole-3-butyric acid (IBA) (0.2, 0.5, and 1 mg/L), BAP (1, 2, 3, and 4 mg/L) + NAA (0.2, 0.5, and 1 mg/L), and TDZ (1, 2, 3, and 4 mg/L) + NAA (0.2, 0.5, and 1 mg/L) were used in the second set of experiments. At the end of the second set of experiments, the greatest callus regeneration ratios were obtained due to the combinations including BAP and IBA as well as the low-dose TDZ- (especially 1 mg/L) and NAA- (0.2, 0.5, 1 mg/L) combinations. Regarding callus fresh weights, TDZ + NAA combinations were found to be more successful. The greatest callus fresh weight (12.7 ±0.4 g) was obtained from MS medium supplemented with 2 mg/L TDZ and 0.2 mg/L NAA.

Fava bean is an extremely important legume and serves immense potential to function as an ingredient as pulse proteins in human diet. Bearing the proficiency of yielding magnanimous amount of functional and nutritional ingredients, this bean deserves to replace any other leguminous crop too. The instability of fava bean in its yield makes breeding for crop improvement difficult, and its high susceptibility to a number of abiotic and biotic stresses additionally results in unstable yields. The self-incompatibility leads to the formation of a limited genetic pool and shows a slow progress in breeding. The plant is highly recalcitrant, making it an onerous task to micropropagate or regenerate fava beans under in vitro conditions. Another fly in the ointment is the release of phenolic compounds by the plant. There are several endeavours that have been made to establish in vitro regeneration, protoplast culture, and genetic transformation and to genetically improve this plant. Nonetheless there are a number of promising cutting-edge technologies that are yet to be harnessed in order to ensure its comprehensive and sustainable genetic improvement. The in vitro-based technologies of this legume and its untraveled path in the plant tissue culture-mediated approaches can assist further genetic manipulation of this plant species in a smoother manner and at an exponential rate. Creation of a single review comprising all the updates and genetic advancements in fava bean is an absolute necessity of the hour. Thus, the importance of this review remains at the peak as it covers a vast range of information, starting from the basic description to the utmost modern stages of advancement in the selected crop. Overall interpretation of the review is aimed at encouraging readers to focus on almost all possible dimensions of international research, already executed, and is being executed in fava bean, thereby helping to understand the demand and advantages of the crop, even at the molecular level. Key points • Fava bean, commonly known as "poor man's meat", is a protein-rich legume with multiple nutritional and pharmacological benefits. • Its highly recalcitrant response makes in vitro interventions quite challenging for its genetic improvement. • This review delves into biotechnological interventions for its advancements to date and highlights major hurdles and potential research solutions to ensure comprehensive genetic improvement. [ABSTRACT FROM AUTHOR]

Rheum ribes L. is a perennial wild species. Young shoots and flower bunches are freshly consumed, and root and rhizomes are generally used for medicinal purposes. The aim of the present study was to improve the callus proliferation protocol for R. ribes L. under in vitro conditions. For callus induction, hypocotyl explants taken from 14-day old plantlets germinated in Murashige and Skoog (MS) media were cultured in MS media with 9 plant growth regulator (PGR) combinations containing 6-benzylaminopurine (BAP) (2, 3, and 4 mg/L) + naphthylacetic acid (NAA) (0.1, 0.5, and 1 mg/L). Then, for callus proliferation, 4 PGR combinations containing NAA (0.2 mg/L) + thidiazuron (TDZ) (0.5, 1, 2, and 3 mg) were used in the first set of experiments, and 36 PGR combinations containing BAP (1, 2, 3, and 4 mg/L) + indole-3-butyric acid (IBA) (0.2, 0.5, and 1 mg/L), BAP (1, 2, 3, and 4 mg/L) + NAA (0.2, 0.5, and 1 mg/L), and TDZ (1, 2, 3, and 4 mg/L) + NAA (0.2, 0.5, and 1 mg/L) were used in the second set of experiments. At the end of the second set of experiments, the greatest callus regeneration ratios were obtained due to the combinations including BAP and IBA as well as the low-dose TDZ- (especially 1 mg/L) and NAA- (0.2, 0.5, 1 mg/L) combinations. Regarding callus fresh weights, TDZ + NAA combinations were found to be more successful. The greatest callus fresh weight (12.7 ±0.4 g) was obtained from MS medium supplemented with 2 mg/L TDZ and 0.2 mg/L NAA. [ABSTRACT FROM AUTHOR]

The problems encountered in the production of the lentisk trees, which are one of the important endemic plants of our country have led to the use of biotechnological methods. In this research for this purpose, the TCL (Thin Cell Layer) technique was considered to investigate of in vitro regeneration potential of expants used for production of lentisk. Firstly, the leaf, node and stem parts of the plant were cut by TCL technique and these explants had been cultured in semi-solid MS media supplemented with 2,4-D and KIN at different concentrations (0.1, 0.2, 0.5 and 1.0 mg/L). The highest callus formation percentage was 100% in transverse stem layers and longitudinal node in MS media including 1 mg/L 2,4-D and 1 mg/L KIN. The lowest callus regeneration ratios were found as 26.67% for three explant types (transverse leaf, transverse stem, longitudinal node). Due to the high rate of darkening in regenerated calli, these were transferred primarily to semi-solid media containing different antioxidants (ascorbic acid, citric acid, PVP, active charcole) and after that cultured in liquid media containing different plant growth regulators (IAA, KIN and BAP) to induced somatic embryogenesis. Later, the calli were encapsulated to prevent darkening and the nurse technique was applied with Aloe vera L. and Gossypium hirsutum L. calli as a different application. As a result of all these trials, somatic embryogenesis didn't occur, but darkening ratio was reduced to 6.67%. [ABSTRACT FROM AUTHOR]

In vitro raised plantlets were obtained from nodal tissue through direct organogenesis and they served as donor plants for the collection of leaf explants. Leaf explants were inoculated on Murashige and Skoog's medium with different concentrations of 2,4-Dichlorophenoxyacetic acid (2,4-D). Hundred percent callogenesis was observed on medium supplemented with 5 mg l−1 2,4-D. For the multiplication of cells (proliferation), calli were transferred to either basal medium or media containing different types and concentrations of cytokinins. Proliferation was observed maximum on media containing 0.5 mg l−1 BAP or 1.0 mg l−1 BAP. Shoot differentiation from calli took place on media supplemented with BAP in combinations with TDZ or ZN. Initiation of organogenesis was observed in calli within two weeks of their subculture on differentiation medium. Shoot differentiation was maximum, when calli proliferated on medium having 1 mg l−1 BAP were transferred to the medium containing 1 mg l−1 BAP and 0.5 mg l−1 TDZ. Organogenic responses after four weeks of subculture on above differentiation medium were as such: number of shoots per explants (25.33 ± 0.88), number of shoots per calli replicate (3.67 ± 0.33) and maximum shoot length (3.43 ± 0.20 cm). Medium supplemented with 2.5 mg l−1 NAA was most responsive for rooting of shoots (54.16 ± 1.39%). About 62.5% plantlets survived after hardening and 54.17% plantlets got acclimatized. All acclimatized plants were transferred to field condition successfully. Formation of unipolar shoots and their multicellular attachment with callus were observed by scanning electron microscopy. To confirm the genetic fidelity of micropropagated plants, five micropropagated plants derived from different leaf explants and two mother plants (randomly selected from micropropagated plants raised from nodal explants) were subjected to molecular analysis. The genetic fidelity of in vitro regenerated plants was assessed by using SCoT and ISSR molecular markers. 12.5% polymorphism was reported in both studies, which may be due to callus mediated regeneration of shoots. Key message: Indirect organogenesis from leaf tissues of Spondias pinnata can be used both for conservation as well as the improvement of plants. [ABSTRACT FROM AUTHOR]

This report focuses on the crucial role of lipids and starch metabolism in the growth and ultrastructure of the cell wall (CW) in rice calli. Non-habituated calli (NH-calli) were isolated from a rice embryo-derived callus culture supplemented with 2,4-D (2,4-Dichlorophenoxyacetic acid); however, the habituated calli (H-calli) were isolated from NH ones and cultured subsequently on exogenous hormone-free medium. The phenotypes of the H- and NH-calli differed significantly between each other as reported in our recently published work. The H-calli CW was thinner, the endoplasmic reticulum (ER), mitochondria (MT) and starch grains (SGs) were less in the H-calli. The intracellular free fatty acids (FAs) and starch content were much less in the H- if compared to NH-calli. By microarray analysis, the differentially expressed genes (DEGs) related to primary and secondary metabolism, lipid and starch metabolism were down-regulated in H- vs NH-calli. The response at the transcriptomic level involved in these changes, as well as the corresponding variations in cytology and FAs metabolism were also accurately elucidated herein. The study reveals that the decreased levels of lipids and starch in the H-calli are mainly attributable to their (H-calli) elevated consumption, in the absence of exogenous hormonal supply to ensure the demand in energy of the calli to survive and maintain an optimum growth and development with the available resource of energy (starch and lipids). This report displays as well the slowdown of the H-calli metabolism, in the absence of exogenous growth regulators supply, if compared to that of the NH-ones. Key message: Lipids and starch decreased in the H-calli is mainly attributable to their elevated consumption to survive and maintain an optimal growth and development with the available resource of energy. [ABSTRACT FROM AUTHOR]

To establish a dynamic and fine suspension culture, four different methods of tomato cell suspension culture were compared. Hypocotyl explants of the tomato cultivar Jina were used for callus induction on Murashige and Skoog (MS) Medium supplemented with three different phytohormone combinations. Then, one gram of each type of calli was transferred to 50 mL of liquid MS medium with four combinations of auxins and cytokinins to produce cell suspensions. The growth rate, judged by cell turbidity, cell fresh weight, and cell viability was evaluated. The best suspension culture was obtained by using friable calli formed on MS medium containing 1 mg L-1 NAA and 0.1 mg L-1 kinetin, transferred to the liquid MS supplemented with 2 mg L-1 NAA, 0.2 mg L-1 2, 4-D and 0.2 mg L-1 zeatin. [ABSTRACT FROM AUTHOR]

Summary: Root‐knot nematodes (RKNs; Meloidogyne spp.) induce new post‐embryogenic organs within the roots (galls) where they stablish and differentiate nematode feeding cells, giant cells (GCs). The developmental programmes and functional genes involved remain poorly defined.Arabidopsis root apical meristem (RAM), lateral root (LR) and callus marker lines, SHORT‐ROOT/SHR, SCARECROW/SCR, SCHIZORIZA/SCZ, WUSCHEL‐RELATED‐HOMEOBOX‐5/WOX5, AUXIN‐RESPONSIVE‐FACTOR‐5/ARF5, ARABIDOPSIS‐HISTIDINE PHOSPHOTRANSFER‐PROTEIN‐6/AHP6, GATA‐TRANSCRIPTION FACTOR‐23/GATA23 and S‐PHASE‐KINASE‐ASSOCIATED‐PROTEIN2B/SKP2B, were analysed for nematode‐dependent expression. Their corresponding loss‐of‐function lines, including those for LR upstream regulators, SOLITARY ROOT/SLR/IAA14, BONDELOS/BDL/IAA12 and INDOLE‐3‐ACETIC‐ACID‐INDUCIBLE‐28/IAA28, were tested for RKN resistance/tolerance.LR genes, for example ARF5 (key factor for root stem‐cell niche regeneration), GATA23 (which specifies pluripotent founder cells) and AHP6 (cytokinin‐signalling‐inhibitor regulating pericycle cell‐divisions orientation), show a crucial function during gall formation. RKNs do not compromise the number of founder cells or LR primordia but locally induce gall formation possibly by tuning the auxin/cytokinin balance in which AHP6 might be necessary. Key RAM marker genes were induced and functional in galls. Therefore, the activation of plant developmental programmes promoting transient‐pluripotency/stemness leads to the generation of quiescent‐centre and meristematic‐like cell identities within the vascular cylinder of galls.Nematodes enlist developmental pathways of new organogenesis and/or root regeneration in the vascular cells of galls. This should determine meristematic cell identities with sufficient transient pluripotency for gall organogenesis. [ABSTRACT FROM AUTHOR]

DUE to increasing importance of roses in producing secondary products including essential oils and pharmaceutical compounds as phenolics, several protocols were developed from different explants of roses. For that purpose, an efficient callus initiation procedure for Rosa damascena Mill. cultivated in Taif, KSA was developed by using petal explant cultures supplemented with nine elicitors for improving phenolics content. As well, related genetic expression variation of the elicited calli was studied by biochemical (protein and isozyme) analyses. Except for rose and geranium oils, seven elicitors increased the phenolics content in R. damascena calli compared with the control. High qualitative differences were observed in their banding patterns reflecting biochemical aberrations in treated calli. A dendrogram, based on UPGMA method of cluster analysis, upheld the notable fluctuations in phenolics content and gene expression occurred by different elicitors. [ABSTRACT FROM AUTHOR]

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Jerusalem artichoke is one of the most widely used plant groups in industrial and medicinal purposes and has been extensively used for inulin production. This plant is well known as a perennial plant and its tubers are rich in inulin. This study aimed to establish an applicable protocol for calli induction and production using different explants of Jerusalem artichoke cultivars (Balady, Fuza and Alba). Leaf, stem and root explants derived from in vitro growing plantlets were cultured on MS medium augmented with different combinations of naphthalene acetic acid (NAA) and benzyl adenine (BA). The highest percentage of calli induction (100%) was recorded with all tested media except MS free growth regulators medium and MS fortified with 1 mg L-1 BA. The maximum value of calli fresh weight was obtained by culturing stem explants on MS supplemented with 2 mg L-1 NAA and 1 mg L-1 BA for all cultivars. Calli fresh weights increased weekly reaching the maximum value at the eighth week for all used explants cultured on the best medium for calli initiation. ‘Balady’ recorded the best results for calli induction and production as compared with ‘Fuza’ and ‘Alba’. Stem explant was superior to root and leaf explants for all examined cultivars. The most suitable medium was MS medium fortified with 2 mg L-1 NAA and 1 mg L-1 BA compared to the other tested media for inducing calli and to enhance calli production from stem explant for all cultivars. [ABSTRACT FROM AUTHOR]

The morphogenic callus (MC) of Fagopyrum tataricum contains a large amount of flavonoids, especially rutin, and exhibits a high level of antioxidant activity. A non-morphogenic callus (NC) may appear on the surface of the MC after two to three years of cultivation and is then subjected to a consistently high level of oxidative stress. The elucidation of the molecular background of this instability is essential for gaining a better understanding of the somaclonal variation mechanisms in tissue cultures that have different morphogenic potentials. Thus, in this study we show that continuous oxidative stress in a NC might be connected with a rapid senescence process and as a result, in the upregulation of the genes that are connected with the telomere complexity, ethylene biosynthesis and the expression of DNA methyltransferases. Moreover, we analysed the presence of the hydroxyproline-rich glycoproteins in the calli and demonstrated the differences between the MC and NC. The LM2 antibody can be useful as a marker of the cells in the MC that are embryogenically determined, while the MAC207 antibody seems to be a positive marker of a MC as its signal was absent in the NC. This study also provides the first report on the effect of trichostatin A on the DNA methyltransferases and demethylases in a MC. Key message: LM2 can be used as a marker of the embryogenically determined cells. Overproduction of ethylene is present in the NC. [ABSTRACT FROM AUTHOR]

One of the principal limitations in application of cell and tissue culture techniques for improving crop plants resistance to adverse environmental stresses, such as high salinity, is insufficient knowledge of cellular and molecular-genetic basics of this type of tolerance. This approach will provide opportunity to identify new salt-tolerant cultivars, which arisen from somatic mutations increasing the pool of salttolerant breeding lines. It is well known that salinity causes sharp reduction in bean productivity and thus significant losses in quality and quantity of products derived from it. Choosing salt-tolerant plant genotypes for cultivation may solve this problem. The aim of this study is to trace tolerance and accompanying changes in lectin accumulation in calli of common bean (Phaseolus vulgaris L.) grown under the conditions of tissue culture. In our experiment, induction of certain common bean cultivars by in vitro cell cultures have been optimized. The most appropriate composition of the nutrient media for the induction of callusogenesis have been established together with the cultivars possessing high callus-forming ability. We have not observed strict correlation between callus-forming propensity and morphogenic calli generation. However, we noticed that under-passaging in selective conditions calli leads to gradual growth decline and browning, as well as slow growth and in some cases death of cell cultures. Nevertheless, we have identified common bean cultivars with average salt tolerance and high propensity to callusogenesis for use as a starting material for breeding. Differences in lectin content between morphogenic and non-morphogenic calli let us suggest that lectin content depends on hormonal composition of the nutrition medium since morphogenic type of callus was formed when the media contained NAA and low concentrations of 2,4-D. The cultivars used in this study have demonstrated moderate salt tolerance and high callusogenesis efficiency thus regarded as suitable material to breeding for salt tolerance. [ABSTRACT FROM AUTHOR]

The production and commercialization of natural antioxidants is gaining increasing importance due to their wide range of biological effects and applications. In vitro cell culture is a valuable source of plant bioactive compounds, especially those highly dependent on environmental factors. Nonetheless, research on the accumulation in plant cultured cells of water-soluble antioxidant vitamins, such as the ascorbic acid (AsA), is very limited. Tomato fruits are a main dietary source of vitamin C and in this work, we explored the potential of in vitro cultured cells for AsA accumulation. Specifically, using a full factorial design, we examined the effect of the source explant, the time in tissue culture and the genetic difference present in two Introgression Line (IL7-3 and IL12-4) that harbor Quantitative Trait Loci (QTLs) for ascorbic acid in fruits. Moreover, we performed an expression analysis of genes involved in AsA metabolism to highlight the molecular mechanisms that can account for the difference between fruit explants and calli. Our work indicated that cultured tomato cells accumulate AsA well beyond the amount present in fruits and that the three factors under investigation and their interaction significantly influence AsA accumulation. The time in tissue culture is the main single factor and, different from the expectations for secondary metabolites, explants from unripe, mature green fruits provided the highest increase in AsA. Moreover, in controlled conditions the genetic differences between the ILs and the control genotype are less relevant for calli cultivated for longer time. Our work showed the potential of tomato cell culture to produce AsA and prompt further refinements towards its possible large-scale exploitation.

2-(2-Phenylethyl)chromones are main classes of aromatic compounds isolated from agarwood, the resinous portion derived from Aquilaria sinensis used as traditional medicine, incense and perfume. However, the mechanisms of biosynthesis of 2-(2-phenylethyl)chromones and agarwood formation are still unknown. Previous studies indicated that 6,7-dimethoxy-2-[2-(4′-methoxyphenyl)ethyl]chromone (AH8) and 6,7-dimethoxy-2-(2-phenylethyl)chromone (AH6) were more sensitive to salt stress and can be used as markers to analyze the production of 2-(2-phenylethyl)chromones. In this study, the content of AH6 and AH8 was analyzed by an AB Sciex 5500 Qtrap mass spectrometer and the congeners of 2-(2-phenylethyl)chromones were detected by LC-DAD-IT-TOF-MS system in A. sinensis calli under various abiotic stresses and hormone treatments. Among the abiotic stresses, salt and drought stress remarkably increased the production of AH6 and AH8, and could induce 41 and 23 species of 2-(2-phenylethyl)chromones, respectively. However, cold treatment has little influence on the production of 2-(2-phenylethyl)chromones. Ion stresses induced by KNO3, CaCl2, and (NH4)2SO4 also significantly increased the content of AH6 and AH8, and could induced 14, 18, and 14 congeners of 2-(2-phenylethyl)chromones, respectively. Exogenous hormones including MeJA, SA and ABA induced a small amount of 2-(2-phenylethyl)chromones in calli within 20 days, while 6, 5, and 7 species of 2-(2-phenylethyl)chromones were detected after MeJA, SA and ABA treatment, respectively. Our study would provide solid basis for further research on biosynthesis of 2-(2-phenylethyl)chromones and agarwood formation. [ABSTRACT FROM AUTHOR]

Color is a key factor for fruit commercial value. 5-Aminolevulinic acid (5-ALA), as an eco-friendly plant growth regulator, shows an attractively promotive effect on plant secondary metabolism, especially for fruit coloration. Brassinosteroids (BRs) can also improve plant flavonoid biosynthesis. No information is now available on the relationship between 5-ALA and BR. Here, we found that 1.5 mg L−1 24-epibrassinolide (24-EBL) promoted 50 mg L−1 5-ALA-induced anthocyanin accumulation, while, brassinazole (Brz) significantly inhibited the 5-ALA-induced flavonoid accumulation. HPLC analysis further showed that the inductive effects of 5-ALA on the accumulation of cyanidin-3-galactoside, quercetin-3-galactoside, quercetin and kaempferol were elevated by 24-EBL, but impaired by Brz. These results suggest that brassinolide biosynthesis might involve in 5-ALA-induced flavonoid accumulation. Gene expression analysis showed that 5-ALA and 5-ALA + 24-EBL induced the expression of regulatory genes MdMYB10, MdMYB9, MdbHLH3 and MdbHLH33. These two treatments also up-regulated the structural gene expressions of anthocyanin biosynthesis and transportation, including MdCHS, MdF3′H, MdDFR, MdANS, MdUFGT, MdGST and MdMATE, as well as flavonol biosynthetic gene MdFLS. But Brz decreased 5-ALA-induced up-regulation of these genes. In addition, 5-ALA also induced the expression of MdBRI1, MdBAK1 and MdBZR1, which are involved in brassinolide signal transduction. These results indicate that 24-EBL enhances 5-ALA-promoted expression of genes related to flavonoid biosynthesis and brassinolide signal transduction, while Brz exhibits the opposite effects. Taken together, we propose that 24-EBL is involved in 5-ALA-induced anthocyanin and flavonol accumulation in calli of apples. Our results provide new insights into 5-ALA-induced fruit coloration. [ABSTRACT FROM AUTHOR]

In this study, an embryogenic callus induction and proliferation system for Hippeastrum vittatum was established, with the tender bulbs as expiants. And then the sensitivity of the expiants and calli to kanamycin and hygromycin was evaluated. The results suggested that the embryogenic calli were induced from tender bulbs and cultured in Murashige and Skoog (MS) medium supplemented with 0.5 mg/L N-phenyl-N'-l, 2,3-thiadiazol-5-ylurea (TDZ), 1.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D), and 30 g/L sucrose (pH5.8) in the dark at 25 ± 1°C. Further study of the influence of kanamycin and hygromycin on callus induction and multiplication showed that, the lethal doses of kanamycin and hygromycin to bulb expiants were 100 and 30 mg/L, respectively. All expiants of H. vittatum died on the medium supplemented with 100 mg/L kanamycin or 30 mg/L hygromycin at the induction stage, and callus proliferation was completely inhibited by 100 mg/L kanamycin or 25 mg/L hygromycin, and all the calli died at last. These results will provide important reference for further studies of transgenic H. vittatum. [ABSTRACT FROM AUTHOR]

Coconut is a very important plant for the livelihood of people in tropical countries. It is also used as an icon of tropical region. Coconut fruit is very heavy and can cause injuries if the fruit falls down and hits somebody who happens to be underneath a coconut tree. In order to avoid the accident,  the coconut fruits have to be regularly cut off. Coconut tree originated from in vitro cultured endosperm is a triploid plant that produces seedless fruit (without endocarp). Coconut fruit without endocarp is not heavy. The objective of this study was to investigate plant regeneration of fresh and in vitro cultured coconut endosperms. The fresh and developed in vitro cultured coconut endosperms were observed using histological analysis. Solid endosperm of seven month-old postanthesis coconut from “Samoan Dwarf†cultivar was freshly picked up and cultured in vitro on modified Branton & Blake formula. Histological study of fresh coconut endosperm showed that the endosperm consisted of parenchyma cells, which were relatively uniform in shape and size, with some nuclei consisted of 1 – 5 nucleoli. Three month-old calli of in vitro grown coconut endosperm in semi solid media showed that its cells varied in shape and size, characterized by high nucleus to cytoplasm ratio, high starch, protein and lipid contents which underwent many divisions. Seven month-old calli of in vitro grown coconut endosperm in liquid media showed embryogenic cells which resembled proembryos. Fourteen month-old bud-like structure of coconut endosperm in semi solid media showed a meristematic layer, tunica-corpus structure, cortex-like region and tracheids of xylem. These results indicated that the bud-like structure was an early stage of shoot bud formation in coconut endosperm. This is the first report of early stage of shoot bud formation occurring on coconut endosperm cultured in vitro.

Lopezia racemosa Cav. is a plant used in Mexican traditional medicine to heal inflammatory diseases. From this plant we isolated the novel compound 6-O-palmitoyl- 3-O-β-D-glucopyranosylcampesterol (1) and 6-O-palmitoyl-3-O-β-D-glucopyranosyl-β-sitosterol (2), previously reported to have cytotoxic activity on several cancer cell lines. We evaluated the anti-inflammatory activity of 1 in vivo by mouse ear edema induced with 12-O-tetradecanoylphorbol-13-acetate (TPA) and 57.14% inhibition was observed. The aim of our study was to obtain callus cultures derived from this plant species with the ability to produce the compounds of interest. Callus cultures were initiated on MS basal medium amended with variable amounts of naphthaleneacetic acid (NAA), or 2,4-dichlorophenoxyacetic acid (2,4-D), combined or not with 6-benzylaminopurine (BAP). Ten treatments with these growth regulators were carried out, using in vitro germinated seedlings as source of three different explants: hypocotyl, stem node, and leaf. Highest yield of 1 was observed on callus derived from leaf explants growing in medium containing 1.0 mg/L 2,4-D and 0.5 mg/L BAP. Selected callus lines produced less 1 than wild plants but the in vitro cultured seedlings showed higher production. So we conclude that it could be attractive to further investigate their metabolic potential.

Garcinia brasiliensis, popularly known as Bacupari, is native to the Amazon and commonly used in folk medicine for its therapeutic properties. This plant is rich in bioactive compounds like benzophenones. However, there are no works about the in vitro establishment and achievement of secondary metabolites in this plant. Thus, the aim of this work was to determine the growth curve and to perform the biochemical and phytochemical analyses in calli obtained from the procambium segments of Bacupari. The growth curve of calli followed a sigmoidal pattern, with four distinct phases (lag, exponential, linear, deceleration). Total soluble sugars were higher on the inoculation day and the reducing sugars on the 20 th day. Amino acids increased from the 60 th day up to the stabilization on the 120 th day. The protein content varied, but it seemed to be related to the amino acids metabolism. The phytochemical screening showed the presence of phenolic and flavonoid compounds in the calli and the HPLC analysis allowed the identification of Fukugetin, Guttiferone A and 7-epiclusianone.

To find the characteristics of somatic embryogenesis of orchids and elucidate the mechanism, we had previously established an efficient plant regeneration system via somatic embryogenesis in Dendrobium candidum Wall ex Lindl. In this study, a detailed cytological investigation was carried out on the initiation and developmental process of somatic embryogenesis. Based on our observations, the somatic embryogenesis in D. candidum originated from the transition of an embryonic callus cell to the initial somatic embryo cell, and the somatic embryos initiated from those cells. During the transition process, condensation and devacuolation successively occurred in the cytoplasm of the embryonic callus cells, giving rise to the formation of a typical initial somatic embryo cell with dense cytoplasm and a clear nucleus. One of the two pathways in somatic embryogenesis is the single-cell-derived somatic embryo which is generated from an inner initial somatic embryo cell in embryonic callus and develops into a globular somatic embryo in a way similar to zygotic embryogenesis and then keeps developing into a protocorm-like body (PLB). The other is a multiple-cell-derived somatic embryo which is generated from peripheral grouped initial somatic cells in embryonic calli and directly forms globular embryo or multicellular somatic proembryo, lacking the typical early stages of embryogenesis. Both pathways were observed in the somatic embryogenesis system, indicating that the culture system in D. candidum can be a useful tool for investigating the mechanisms underlying orchid embryogenesis. [ABSTRACT FROM AUTHOR]

While many redox enzymes are nowadays available for synthetic applications, the toolbox of ene-reductases is still limited. Consequently, the screening for these enzymes from diverse sources in the search of new biocatalyst suitable for green chemistry approaches is needed. Among 13 plant tissue cultures, Medicago sativa and Tessaria absinthioides calli, as well as Capsicum annuum hairy roots, were selected due to their ability to hydrogenate the C C double bond of the model substrate 2-cyclohexene-1-one. The three axenic plant cultures showed more preference toward highly activated molecules such as nitrostyrene and maleimide rather than the classical substrates of the well-known Old Yellow Enzymes, resembling the skills of the NAD(P)H-dependent flavin-independent enzymes. When the three biocatalytic systems were applied in the reduction of chalcones, T. absinthioides showed high chemoselectivity toward the C C double bond whereas the other two demonstrated abilities to biohydrogenate the C C double bounds and the carbonyl groups in a sequential fashion. [ABSTRACT FROM AUTHOR]

The aim of the present research was to determinate the type of tissue present in a sample obtained from an embryogenic callus of Coco Cumbé Palm, Parajubaea cocoides Burret and somatic embryos on globular stage. For this purpose, in order to the protocol established in the histotechnology analysis procedure manual of Patology Service from "Carlos Andrade Marín" Hospital (HCAM, according its acronyms in Spanish) was proceeded located in Quito, Pichincha, Ecuador. The results showed that the embryogenic calli are conformed by meristematic cells and although present cells with starch granules near to embryogenic cells, indicating embryos formation. In somatic embryos case, was verified that these were in globular stage, because of these showed a characteristic structure of the first development stage, named protoderm. Therefore was determined that calli present distinctive characteristics of embryogenic competence and viability of the somatic embryos. [ABSTRACT FROM AUTHOR]

This study characterized the antagonistic activities of five fungal endophytes ( Aspergillus calidoustous BTF07, Penicillium citrinum BTF08, Trichoderma asperellum T2, Diaporthe phaseolorum WAA02, Diaporthe phaseolorum MIF01) and evaluated their endophyte-host compatibility with the host plant (oil palm). The antifungal activities of the endophytes towards Ganoderma boninense (GB) were first established using the dual culture test, revealing antagonistic nature of endophytes via production of non-volatiles, volatiles and competitive exclusion. Endophyte-host compatibility was then assessed using a simple but rapid endophyte-calli dual-culture assay, and results validated using endophyte-ramet test. Results revealed that endophytes elicited different responses in oil palm calli. BTF08 had growth promoting effects towards the host tissues with the highest calli weight (1013 mg) obtained, while BTF07 appeared to inhibit calli (1006 mg) leading to browning and necrosis. This endophyte-calli test also revealed the influence of calli on endophyte growth. Isolates BTF08 and GB benefited from host association, with increased radial growth (2.36 cm and 2.31 cm, respectively) compared to growth in the absence of calli (2.10 cm and 2.15 cm, respectively). Endophytes and GB were also isolated from host tissues, suggesting compatibility and ability to colonize host tissues (root, stem, leaf). This suggested the reliability of the endophyte-calli test as a rapid assay to provide an insight on the endophyte-host compatibility. [ABSTRACT FROM AUTHOR]

Friable calli of Polygonum multiflorum Thunb have been induced in MS medium supplemented with 6-benzylaminopurine (6-BA) and kinetin (KT). Suspension cultures were initiated from friable calli by inoculating calli in liquid MS medium in shake flasks in the dark and 25 °C on an orbital shaker at 100 rpm. The maximum dry weight (DW, 7.85 g/L) and 2,3,5,4′-tetrahydroxystilbene-2-O-β-D-glycoside (THSG, 56.39 mg/L) of suspension cells was obtained in MS medium after 16 days culture. Both methyl jasmonate (MeJA) and salicylic acid (SA) could increase THSG production. The most appropriate concentration of MeJA was 100 μmol/L in MS medium, in which concentration THSG content reached the maximum value of 147.79 mg/L, which represented a 162.36% increase compared to that of the control (56.33 mg/L). The most appropriate concentration of SA was 125 μmol/L in MS medium, at which concentration THSG content reached its maximum value of 116.43 mg/L, a 106.69% increase compared to that of the control (56.33 mg/L).

Valeriana pyramidalis Kunth, es una planta nativa, popularmente utilizada por su actividad calmante en el tratamiento de problemas nerviosos y cardíacos, además de enfermedades y trastornos como migraña e insomnio. En la presente investigación se planteó el establecimiento de suspensiones celulares de V. pyramidalis Kunth, como un estudio base para una futura cuantificación de sus metabolitos secundarios. Tras estandarizar un protocolo de desinfección y obtener callos friables, se establecieron suspensiones celulares y se obtuvo con éxito las curvas de crecimiento celular.

Restrepia is a small Pleurothallid genus, comprising 57 species, 44 of which were discovered since 1970. These species are indigenous to Central and South America, where their montane forest habitats are under increasing pressure from changes in land use. With resulting increasingly fragmented habitats and dwindling numbers, the pollination systems of obligate out-breeding genera, such as Restrepia , may no longer function efficiently which could potentially lead to their extinction. As such, the main aim of the current study was to perform an in-depth investigation of floral structures in the genus, using SEM and photographic technology to formulate a putative pollination mechanism for these species. The floral micromorphology of dorsal sepal and lateral petal osmophores, synsepal, labellum, cirrhi and calli were investigated by scanning electron microscopy (SEM), macro-photography and quantitative analyses of some floral proportions. The secretory nature of the labellum, synsepal and osmophore papillae were established and the calli were shown to possess a unique papillate, non-secretory structure. A pollination mechanism for the genus was proposed which includes the role of the scent trails produced by the osmophores and the ‘trapping’ role of the cirrhi. A ‘functional fit’ between the flower and the pollinator is suggested. In conclusion, we consider Restrepia to represent a non-nectar rewarding and ‘deceptive’ orchid genus and that this pollination mechanism may be directly linked to the breeding system (gametophytic self-incompatibility) in this genus. [ABSTRACT FROM AUTHOR]

Sphaeralcea angustifolia is used in Mexican traditional medicine to treat inflammatory processes. SCopoletin (SC), TOmentin (TO), and sphaeralcic acid (SA) were reported as the main anti-inflammatory compounds in this species. The aim of this study was to establish in vitro conditions for the development of calli and cell suspension cultures that are the producers of these active compounds. Callus cultures of plant leaf explants were set up using different auxin levels of α-naphthalene acetic acid (NAA) in combination with a constant concentration (0.1 mg L) of Kinetin (Kn) in Murashige and Skoog (MS) medium. Optimal combinations for callus induction were 1.0 and 2.0 mg L of NAA. SC, TO, and SA were not detected in callus tissues. Employing a 4 % inoculum in fresh biomass, cell suspension was established from friable callus with 1.0 mg L of NAA in combination with 0.1 mg L of Kn in MS liquid medium (27.4 mM nitrate). The cellular suspension synthesized SC and SA, SC was excreted into the culture medium, while SA was excreted into the culture medium and accumulated in biomass. To improve SC and SA production, total nitrate content was reduced in MS medium. On diminishing nitrate content to 2.74 mM, cellular suspension growth was not modified. SC concentration (0.04 %) was 60-fold higher than that detected in the wild plant (0.00067 %), TO was produced (0.096 %), and SA content (0.0036 %) was not improved. SA production in MS medium with 0.274 mM nitrate (0.004 %) was enriched 12-fold (0.0003 %) in relation to that of the wild plant. The anti-inflammatory effects at 5 h of intraperitoneal (i.p.) administration (100 mg per kg BW) of dichloromethane extracts from the medium (42 ± 3 %) and biomass (39 ± 9.3 %) of S. angustifolia cell suspensions cultivated in MS with 2.74 mM nitrate were similar. The effect of the biomass dichloromethane extract was dose dependent with a median Effective Dose (ED) of 137.63 mg per kg BW. [ABSTRACT FROM AUTHOR]