Plant regeneration of southern Adriatic iris by somatic embryogenesis

A simple protocol has been developed for plant regeneration by somatic embryogenesis of Southern Adriatic iris (Iris pseudopallida Trinajstić), an endemic species of the Balkan Peninsula. Somatic embryogenesis was induced in zygotic embryo culture on media supplemented with 2,4-dichlorophenoxy acetic acid (2-10 mgL-1) as the sole plant growth regulator, where both embryogenic calli and somatic embryos were induced. Subsequent decrease of 2,4-D in the media promoted formation of somatic embryos. Developed somatic embryos germinated on medium without growth regulators. The regenerated plantlets had diploid chromosome number. Planted plantlets acclimatized very well under greenhouse and garden conditions.Jednostavan protokol za regeneraciju biljaka procesom somatske embriogeneze razvijen je za južno-jadranski iris (Iris pseudopallida Trinajstić), endemit Balkanskog poluostrva. Somatska embriogeneza je indukovana u kulturi 'zrelih' zigotskih embriona na hranljivoj podlozi obogaćenoj 2,4-dihlorofenoksi sirćetnom kiselinom (2,4-D), (kao jedinim regulatorom rastenja), gde su embriogeni kalus i somatski embrioni formirali. Smanjenje koncentracije 2,4-D u hranljivoj podlozi dovodi do povećanja broja formiranih somatskih embriona. Klijanje somatskih embriona je postignuto na podlozi bez regulatora rastenja. Regenerisane biljčice su imale diploidan broj hromozoma. Aklimatizacija biljaka je bila dobra u uslovima staklenika i bašte.Projekat ministarstva br. 14263

Superoxide dismutase activity and isoenzyme profiles in bulbs of snake's head fritillary in response to cold treatment

The activities and isoenzyme profiles of superoxide dismutase (SOD) in in vitro Fritillaria meleagris bulbs in response to cold treatment (4°C) were investigated. Differences in SOD activity and isoenzyme profiles in bulbs under standard growth conditions, six weeks chilling, as well as seven days after the completion of cold treatment are presented. SOD activity initially decreased but then rapidly increased seven days after cold treatment. Four isoforms of SOD are active under standard and chilling conditions, while three isoforms are presented 7 days after cold treatment. Native gel electrophoresis indicated the presence of mitochondrial and chloroplast localized SODs.Proučavana je aktivnost i izoenzimski profil superoksid dizmutaze (SOD) u lukovicama Fritillaria meleagris gajenim u uslovima in vitro kao odgovor na tretman hlađenja. Prikazane su razlike u aktivnosti SOD kao i izoenzimskim profilima kod lukovica na standardnim uslovima gajenja, posle šest nedelja hlađenja i sedam dana posle završetka tretmana hlađenja. Aktivnost SOD se smanjuje tokom hlađenja dok se značajno povećava sedam dana posle hlađenja. Četiri SOD izoforme su aktivne tokom gajenja u standardnim uslovima i tokom hlađenja dok su tri izoforme detektovane sedam dana posle hladnog tretmana. Nativna gel elektroforeza je pokazala prisustvo aktivnosti mitohondrijalne i hloroplastne SOD.Projekat ministarstva br. 143026

Clonal fidelity of chrysanthemum regenerated from long term cultures

Morphological characteristics of flowers of long term regenerated chrysanthemum, cv. "White Spider", after ten years of micropropagation are investigated. Shoot cultures are established and maintained more than ten years by stem segment culture on MS medium supplemented with BAP and NAA (1.0, 0.1 mgL-1, respectively). Rooting of shoots (100 %) has done on MS medium without hormones and it was very successful after ten years, as well as, after two or eight years of micropropagation. Acclimation of rooted chrysanthemum plantlets at greenhouse conditions was excellent and after appropriate photoperiod "in vitro" plants flowered 90.3 % and have the same flower color, shape and size as mother plants. Flower color changes of "in vitro" plants are observed during another flowering cycle one year after acclimatization. Observed variations of chrysanthemum flowers could be attributed to epigenetic factors.Praćene su morfološke karakteristike cvetova regeneranata cv. "White Spider" posle deset godina gajenja u kulturi in vitro. Izdanci su umnožavani u kulturi segmenata stabla preko aksilarnih pupoljaka na MS podlozi sa α-naftilsirćetnom kiselinom i benzilaminopurinom (0,5 odnosno 1,0 mgL-1), . Praćeno je umnožavanje izdanaka u periodu od devetnaest pasaža (tri godine) kada je došlo do sukcesivnog smanjenja indeksa umnožavanja. Oživljavanje izdanaka je praćeno na MS podlozi bez hormona i bilo je podjednako uspešno posle dve, kao i posle osam i deset godina mikropropagacije. Oživljene biljke hrizantema su uspešno aklimatizovane na uslove staklenika (97 %). Nakon odgovarajućeg fotoperioda, 90,3 % biljaka je cvetalo i imale su identičnu boju cveta matičnim biljkama hrizantema. Promene u boji cveta su uočene kod "in vitro" biljaka u sledećem ciklusu cvetanja, tj. godinu dana posle aklimatizacije. Ove promene bi se mogle pripisati epigenetskim faktorima.nul

Plant regeneration of southern Adriatic iris by somatic embryogenesis

A simple protocol has been developed for plant regeneration by somatic embryogenesis of Southern Adriatic iris (Iris pseudopallida Trinajstić), an endemic species of the Balkan Peninsula. Somatic embryogenesis was induced in zygotic embryo culture on media supplemented with 2,4-dichlorophenoxy acetic acid (2-10 mgL-1) as the sole plant growth regulator, where both embryogenic calli and somatic embryos were induced. Subsequent decrease of 2,4-D in the media promoted formation of somatic embryos. Developed somatic embryos germinated on medium without growth regulators. The regenerated plantlets had diploid chromosome number. Planted plantlets acclimatized very well under greenhouse and garden conditions.Jednostavan protokol za regeneraciju biljaka procesom somatske embriogeneze razvijen je za južno-jadranski iris (Iris pseudopallida Trinajstić), endemit Balkanskog poluostrva. Somatska embriogeneza je indukovana u kulturi 'zrelih' zigotskih embriona na hranljivoj podlozi obogaćenoj 2,4-dihlorofenoksi sirćetnom kiselinom (2,4-D), (kao jedinim regulatorom rastenja), gde su embriogeni kalus i somatski embrioni formirali. Smanjenje koncentracije 2,4-D u hranljivoj podlozi dovodi do povećanja broja formiranih somatskih embriona. Klijanje somatskih embriona je postignuto na podlozi bez regulatora rastenja. Regenerisane biljčice su imale diploidan broj hromozoma. Aklimatizacija biljaka je bila dobra u uslovima staklenika i bašte.Projekat ministarstva br. 14263

Effect of low temperature on rooting rate and carbohydrate content of Fritillaria meleagris bulbs formed in culture in vitro

Projekat ministarstva br. 143026

Foliar application of methyl jasmonate affects impatiens walleriana growth and leaf physiology under drought stress

In this study, the effects of foliar applied methyl jasmonate (MeJA) on drought-stressed Impatiens walleriana growth and leaf physiology parameters: stomatal conductance, chlorophyll, flavonoid, anthocyanin, and nitrogen balance index (NBI), were evaluated. These parameters could serve as indicators of drought tolerance of I. walleriana, a popular horticultural plant worldwide that is very sensitive to drought. The experiment included four treatments: control, drought-stressed plants sprayed with distilled water, drought-stressed plants sprayed with 5 µM MeJA, and drought-stressed plants sprayed with 50 µM MeJA. Foliar spraying with MeJA was performed twice: seven days before and on the day of drought induction. The stressed plant groups were non-irrigated to achieve soil water contents (SWC) of 15 and 5%, while control plants were well-watered throughout the experiment (35–37% SWC). The results of this study showed that drought significantly reduced I. walleriana fresh and dry shoot weight, as well as total leaf area, but did not impact on dry matter content. The foliar application of MeJA improved growth parameters of I. walleriana, depending on the elicitor concentration and drought intensity. Stomatal conductance was slightly reduced at 5% SWC, and foliar applied MeJA at both concentrations. The flavonoid index was slightly reduced at 15 and 5% SWC when 50 µM MeJA was foliar applied, while there were no observed changes in the anthocyanin index in any treatments. The foliar application of 50 µM MeJA increased the chlorophyll index and NBI of I. walleriana at 5% SWC, indicating a contribution of the elicitor to plant drought tolerance at the physiological level. © 2023 The Author(s). Published with license by Taylor & Francis Group, LLC

Clonal fidelity of chrysanthemum regenerated from long term cultures

Morphological characteristics of flowers of long term regenerated chrysanthemum, cv. "White Spider", after ten years of micropropagation are investigated. Shoot cultures are established and maintained more than ten years by stem segment culture on MS medium supplemented with BAP and NAA (1.0, 0.1 mgL-1, respectively). Rooting of shoots (100 %) has done on MS medium without hormones and it was very successful after ten years, as well as, after two or eight years of micropropagation. Acclimation of rooted chrysanthemum plantlets at greenhouse conditions was excellent and after appropriate photoperiod "in vitro" plants flowered 90.3 % and have the same flower color, shape and size as mother plants. Flower color changes of "in vitro" plants are observed during another flowering cycle one year after acclimatization. Observed variations of chrysanthemum flowers could be attributed to epigenetic factors.Praćene su morfološke karakteristike cvetova regeneranata cv. "White Spider" posle deset godina gajenja u kulturi in vitro. Izdanci su umnožavani u kulturi segmenata stabla preko aksilarnih pupoljaka na MS podlozi sa α-naftilsirćetnom kiselinom i benzilaminopurinom (0,5 odnosno 1,0 mgL-1), . Praćeno je umnožavanje izdanaka u periodu od devetnaest pasaža (tri godine) kada je došlo do sukcesivnog smanjenja indeksa umnožavanja. Oživljavanje izdanaka je praćeno na MS podlozi bez hormona i bilo je podjednako uspešno posle dve, kao i posle osam i deset godina mikropropagacije. Oživljene biljke hrizantema su uspešno aklimatizovane na uslove staklenika (97 %). Nakon odgovarajućeg fotoperioda, 90,3 % biljaka je cvetalo i imale su identičnu boju cveta matičnim biljkama hrizantema. Promene u boji cveta su uočene kod "in vitro" biljaka u sledećem ciklusu cvetanja, tj. godinu dana posle aklimatizacije. Ove promene bi se mogle pripisati epigenetskim faktorima.nul

Alterations in Physiological, Biochemical, and Molecular Responses of Impatiens walleriana to Drought by Methyl Jasmonate Foliar Application

Drought stress affects plant growth and development through several mechanisms, including the induction of oxidative stress. To cope with drought, plants have drought tolerance mechanisms at the physiological, biochemical, and molecular levels. In this study, the effects of foliar application of distilled water and methyl jasmonate (MeJA) (5 and 50 µM) on the physiological, biochemical, and molecular responses of Impatiens walleriana during two drought regimes (15 and 5% soil water content, SWC) were investigated. The results showed that plant response depended on the concentration of the elicitor and the stress intensity. The highest chlorophyll and carotenoid contents were observed at 5% SWC in plants pre-treated with 50 µM MeJA, while the MeJA did not have a significant effect on the chlorophyll a/b ratio in drought-stressed plants. Drought-induced formation of hydrogen peroxide and malondialdehyde in plants sprayed with distilled water was significantly reduced in plant leaves pretreated with MeJA. The lower total polyphenol content and antioxidant activity of secondary metabolites in MeJA-pretreated plants were observed. The foliar application of MeJA affected the proline content and antioxidant enzyme activities (superoxide dismutase, peroxidase, and catalase) in plants that suffered from drought. The expression of abscisic acid (ABA) metabolic genes (IwNCED4, IwAAO2, and IwABA8ox3) was the most affected in plants sprayed with 50 µM MeJA, while of the four analyzed aquaporin genes (IwPIP1;4, IwPIP2;2, IwPIP2;7, and IwTIP4;1), the expression of IwPIP1;4 and IwPIP2;7 was strongly induced in drought-stressed plants pre-treated with 50 µM MeJA. The study’s findings demonstrated the significance of MeJA in regulating the gene expression of the ABA metabolic pathway and aquaporins, as well as the considerable alterations in oxidative stress responses of drought-stressed I. walleriana foliar sprayed with MeJA. The results improved our understanding of this horticulture plant’s stress physiology and the field of plant hormones’ interaction network in general. © 2023 by the authors

Phenol Removal Capacity of the Common Duckweed (Lemna minor L.) and Six Phenol-Resistant Bacterial Strains From Its Rhizosphere: In Vitro Evaluation at High Phenol Concentrations

The main topic of this study is the bioremediation potential of the common duckweed, Lemna minor L., and selected rhizospheric bacterial strains in removing phenol from aqueous environments at extremely high initial phenol concentrations. To that end, fluorescence microscopy, MIC tests, biofilm formation, the phenol removal test (4-AAP method), the Salkowski essay, and studies of multiplication rates of sterile and inoculated duckweed in MS medium with phenol (200, 500, 750, and 1000 mg L−1) were conducted. Out of seven bacterial strains, six were identified as epiphytes or endophytes that efficiently removed phenol. The phenol removal experiment showed that the bacteria/duckweed system was more efficient during the first 24 h compared to the sterile duckweed control group. At the end of this experiment, almost 90% of the initial phenol concentration was removed by both groups, respectively. The bacteria stimulated the duckweed multiplication even at a high bacterial population density (>105 CFU mL−1) over a prolonged period of time (14 days). All bacterial strains were sensitive to all the applied antibiotics and formed biofilms in vitro. The dual bacteria/duckweed system, especially the one containing strain 43-Hafnia paralvei C32-106/3, Accession No. MF526939, had a number of characteristics that are advantageous in bioremediation, such as high phenol removal efficiency, biofilm formation, safety (antibiotic sensitivity), and stimulation of duckweed multiplication

Dimethyl sulfoxide improves sensitivity and specificity of RT-PCR and qRT-PCR amplification of low-expressed transgenes

The expression of transgenes in a host plant may be low for a number of reasons. Both low expression and poor specificity of amplification were encountered during analysis of the expression of the Arabidopsis cytokinin oxidase/dehydrogenase (AtCKX1) gene in transgenic Centaurium erythraea. The optimization of the PCR protocol involved a gradient of annealing temperatures, as well as the application of seven PCR enhancers: formamide, dimethyl sulfoxide (DMSO), glycerol, ethylene glycol, trehalose, BSA and Tween-20. The best results for AtCKX1 amplification were obtained at 55.1ºC, with the addition of 5% DMSO. Glycerol and trehalose also improved the sensitivity of amplification, while formamide, ethylene glycol and BSA enhanced only the amplification of control purified targets, but not the transcripts. Tween-20 inhibited PCR. DMSO enhanced AtCKX1 PCR amplification and improved the specificity of qPCR amplification, as well as the assay reproducibility. This work emphasizes the usefulness of additives, which are rarely used for PCR optimization in real-time experiments.Projekat ministarstva br. ON173015 i br. ON17302