Showing papers on "Kinetin published in 2023"

Cretan Dittany (Origanum dictamnus L.), a Valuable Local Endemic Plant: In Vitro Regeneration Potential of Different Type of Explants for Conservation and Sustainable Exploitation

Abstract: Origanum dictamnus L. is a medicinal local endemic to the Island of Crete, Greece. Its propagation through biotechnological tissue culture techniques is essential due to its augmented multi-industrial sector demand. For direct organogenesis, among different culture media variants (MS, Gamborg B5), and cytokinins [6-benzyladenine (BA), kinetin (Kin), 2-isopentenyl adenine (2-iP)], the MS + added with BA (2.2 μM) was the most effective treatment for shoots and roots formation. For indirect organogenesis, all explant types (leaves, petioles, roots) showed a 100% callusing rate after 2 months in all media variants tested; ODK1: 20 μM thidiazuron (TDZ) + 5 μM indole-3-butyric acid (IBA) or ODK2: 0.5 μM kinetin + 5 μM 2,4-dichlorophenoxy acetic acid (2,4-D). The leaves and petiole explants assured a low rate of shoot regeneration (20%) in ODK1. Afterwards, leaf-, petiole-and root-callus derived from both media were transferred to four new media plant growth regulators—free or with BA + IBA + gibberellic acid (GA3). After 10 months from callus transferring, the petiole callus gave rise to roots (20–75%) while the leaf callus exhibited 10–30% shoot or 30% root regeneration. In this study, indirect organogenesis of O. dictamnus was carried out for the first time, thus various organs can be used for plant regeneration, and the developed protocol may be applicable in the horticulture industry.

Investigation of the Effects of the Explant Type and Different Plant Growth Regulators on Micropropagation of Five Mediterranean Salvia spp. Native to Greece

Abstract: Sages are medicinal and aromatic plants that constitute a large pool from which active compounds of great pharmaceutical potential can be derived, while at the same time, they also have ornamental value. The purpose of this study was to develop the micropropagation protocols of Salvia fruticosa, S. officinalis, S. ringens, S. tomentosa, and S. pomifera ssp. pomifera to facilitate their exploitation in the pharmaceutical and floriculture industries. In vitro cultures of S. ringens and S. pomifera ssp. pomifera was studied for the first time. Shoot tips and single node explants from in vitro seedlings were initially cultured on hormone free (Hf)-MS medium, followed by subcultures on MS medium supplemented with 6-benzyladenine (BA) for all species, as well as with zeatin (ZEA), kinetin (KIN), 6-(γ,γ-dimethylallyamino) purine (2iP), or meta-topolin (mT) for S. fruticosa and S. officinalis, at concentrations 0.0 to 3.2 mg L−1, in combination with 0.01 mg L−1 1-naphthaleneacetic acid (NAA). S. officinalis was the most efficient in shoot multiplication of all the studied species. The highest multiplication indices were found using 0.8 mg L−1 BA for S. fruticosa, 0.4 mg L−1 BA, or mT for S. officinalis, and lower than 0.8 mg L−1 BA for the other three species. Hyperhydricity was a problem at the multiplication stage, and was most pronounced in single node explants, increasing in proportion to cytokinin concentration. Microshoots rooted at high percentages (75–85%) on half-strength MS medium with 0.0 or 0.5 mg L−1 Indole-3-butyric acid (IBA), except for those of S. ringens, which rooted best at 1.0–2.0 mg L−1 IBA. Ex vitro acclimatization was highly successful (80–95%) on peat–perlite substrate (1:1 v/v). Thus, the present study resulted in efficient micropropagation protocols for five Mediterranean sage species native to Greece, which will facilitate breeding programs and the promotion of these species in the floriculture and pharmaceutical industries.

Exogenous Kinetin Modulates ROS Homeostasis to Affect Heat Tolerance in Rice Seedlings

Abstract: Heat stress caused by rapidly changing climate warming has become a serious threat to crop growth worldwide. Exogenous cytokinin (CK) kinetin (KT) has been shown to have positive effects in improving salt and drought tolerance in plants. However, the mechanism of KT in heat tolerance in rice is poorly understood. Here, we found that exogenously adequate application of KT improved the heat stress tolerance of rice seedlings, with the best effect observed when the application concentration was 10−9 M. In addition, exogenous application of 10−9 M KT promoted the expression of CK-responsive OsRR genes, reduced membrane damage and reactive oxygen species (ROS) accumulation in rice, and increased the activity of antioxidant enzymes. Meanwhile, exogenous 10−9 M KT treatment significantly enhanced the expression of antioxidant enzymes, heat activation, and defense-related genes. In conclusion, exogenous KT treatment regulates heat tolerance in rice seedlings by modulating the dynamic balance of ROS in plants under heat stress.

Role of Phytohormones in Biomass and Polyphenol Accumulation in Salvia bulleyana In Vitro Culture

Abstract: Salvia bulleyana is a plant native to the Chinese Yunnan Province. This species has been used in traditional Chinese medicine as a substitute for Danshen (the roots of Salvia miltiorrhiza). The aim of our study was to establish an effective system for propagating S. bulleyana shoots to obtain large amounts of material rich in bioactive compounds. Phytohormones were used to regulate shoot growth and regeneration potential and influence plant secondary metabolism. The shoot tips were incubated on a Murashige and Skoog agar medium supplemented with 0.1 or 0.5 mg/L IAA (indole-3-acetic acid) and the cytokinins benzylaminopurine (BAP), meta-topoline (M-T), 6-benzylaminopurine riboside (RBAP), N-benzyl-9-(2-tetrahydropyranyl)-adenine (BPA) or kinetin, (K) at concentrations of 0.5, 1 or 2 mg/L. It was observed that the type and concentration of growth regulator significantly influenced the regeneration potential of S. bulleyana shoots. The highest multiplication rate was obtained when 0.1 mg/L IAA and 2 mg/L BPA were used. Under these conditions, 100% of shoot tips formed buds and almost seven buds/shoot per explant were obtained after five weeks. Meanwhile, the highest biomass was found for shoots growing on a medium supplemented with 0.1 mg/L IAA and 1 mg/L M-T: 1.2 g of fresh weight and 0.17 g of dry weight. However, a medium with 0.1 mg/L IAA and 2 mg/L RBAP was most favorable for bioactive phenolic acid content, with a total polyphenol level (37.7 mg/g dw) 4.5 times higher than in shoots grown on medium without growth regulators (8.23 mg/g dw). Finally, optimal conditions were selected by TOPSIS (technique for order of preference by similarity to the ideal solution); the culture of S. bulleyana grown on an MS medium containing 0.1 mg/L IAA and 1 mg/L M-T was found to be the most efficient for polyphenol accumulation and can be used for the production of medicinally relevant compounds.

Optimization of In Vitro Propagation of Pear (Pyrus communis L.) ‘Pyrodwarf®(S)’ Rootstock

Abstract: Pears are among the most economically important fruits in the world that are grown in all temperate zones. Pyrus communis L., ‘Pyrodwarf®(S)’ rootstock is one of the gene sources used to improve fruit productivity, rootstock resistance, and tolerance to biotic and abiotic stresses. Traditional propagation of P. communis L. is time-consuming and limited by a short growing season and harsh winter conditions. Therefore, in vitro propagation is a suitable alternative. Murashige and Skoog medium (MS) and woody plant medium (WPM) supplemented with different concentrations of 6-benzyladenine (BA) and kinetin (Kin), individually or in combination, were used for in vitro shoot proliferation. Nodal segments were used as explants. MS medium augmented with indole-3-butyric acid (IBA) or indole-3-acetic acid (IAA) was then used for rooting of microshoots. A combination of 2 mg·L−1 BA and 1 mg·L−1 Kin in MS medium resulted in a significant improvement in shoot proliferation. This combination produced the highest number of shoots (4.352 per explant) and leaves (10.02 per explant). The longest shoots (4.045 cm) were obtained in WPM enriched with 1 mg·L−1 BA. However, these shoots were not suitable for multiplication and rooting steps. The largest number of roots (5.50 per microshoot) was obtained on MS medium augmented with IAA at 1 mg·L−1. The produced plantlets were cultivated in pots filled with perlite and cocopeat (in a ratio of 1:3) and acclimatized gradually in a greenhouse, recording an even 90% survival rate.

Species-Specific Secondary Metabolites from Primula veris subsp. veris Obtained In Vitro Adventitious Root Cultures: An Alternative for Sustainable Production

Abstract: Primula veris subsp. veris L. is a perennial herbaceous and medicinal plant species the roots and flowers of which are a source of valuable pharmaceutical raw materials. The plant tissues are used to produce expectorant and diuretic drugs due to their high content of triterpene saponins and phenolic glycosides. Underground roots of P. veris can be obtained only through a destructive process during the plant’s harvesting. In the present study, an in vitro adventitious root production protocol was developed as an alternative way of production, focused on four species-specific secondary metabolites. Root explants were cultured in Murashing & Skoog liquid medium supplemented with 5.4 μM α-naphthaleneacetic acid, 0.5 μM kinetin, L-proline 100 mg/L, and 30 g/L sucrose, in the dark and under agitation. The effect of temperature (10, 15 and 22 °C) on biomass production was investigated. The content of two flavonoid compounds (primeverin and primulaverin), and two main triterpene saponins (primulic acid I and II) were determined after 60 days of culture and compared with 1.5-year-old soil-grown plants. The accumulated content (mg/g DW) of bioactive compounds of in vitro adventitious roots cultured under 22 °C was significantly higher than the other two temperatures of the study, being 9.71 mg/g DW in primulaverin, 0.09 mg/g DW in primeverin, 6.09 mg/g DW in primulic acid I, and 0.51 mg/g DW in primulic acid II. Compared to the soil-grown roots (10.23 mg/g DW primulaverin, 0.28 mg/g DW primeverin, 17.01 mg/g DW primulic acid I, 0.09 mg/g DW primulic acid II), the in vitro grown roots at 22 °C exhibited a 5.67-fold higher content in primulic acid II. However, primulic acid I and primeverin content were approximately three-fold higher in soil-grown roots, while primulaverin content were at similar levels for both in vitro at 22 °C and soil-grown roots. From our results, tissue culture of P. veris subsp. veris could serve not only for propagation but also for production of species-specific secondary metabolites such as primulic acid II through adventitious root cultures. This would therefore limit the uncontrolled collection of this plant from its natural environment and provide natural products free from pesticides in a sustainable way.

Cytokinin-inducible response regulator SlRR6 regulates plant height through gibberellin and auxin pathways in tomato.

Abstract: Plant height is a key agronomic trait regulated by several phytohormones like gibberellins (GAs) and auxin. However, little is known about how cytokinin (CK) participates in this process. Here, we report that SlRR6, a type-A response regulator in CK signaling pathway, positively regulates plant height in tomato. SlRR6 was induced by exogenous kinetin and GA3, but inhibited by indole-3-acetic acid (IAA). Knockout of SlRR6 reduced tomato plant height through shortening internode length, while overexpression of SlRR6 caused higher plant due to increased internode number. Cytological observation of longitudinal stems showed that both knockout and overexpression of SlRR6 generated larger cells, but significantly reduced cell numbers in each internode. Further studies demonstrated that overexpression of SlRR6 enhanced GA accumulation and lowered IAA content, along with expression changes in GA- and IAA-related genes. Exogenous paclobutrazol and IAA treatments restored the increased plant height phenotype in SlRR6-overexpressing lines. Yeast two-hybrid, bimolecular fluorescence complementation and co-immunoprecipitation assays showed that SlRR6 interacts with a small auxin up RNA protein SlSAUR58. Moreover, SlSAUR58-overexpressing plants were dwarf with decreased internode length. Overall, our findings establish SlRR6 as a vital component in the CK signaling, GA, and IAA regulatory network that controls plant height.

Optimization of Morphogenesis and In Vitro Production of Five Hyacinthus orientalis Cultivars

Abstract: To maximize the economic benefits of Hyacinthus orientalis L., the micropropagation and morphogenesis induction of five Hyacinthus cultivars were investigated under four different concentrations of indole acetic acid (IAA) with two cytokinins, benzyl adenine (BA), or kinetin (Kin). Days for morphogenesis initiation and shoot formation in the red cultivars were fewer than in the blue and white cultivars. Blue cultivars showed an increase in fresh weight and chlorophyll content under either BA or Kin, but they showed an increase in shoot height in the BA treatments only. IAA at 1.5 mg/L caused a time reduction in days for morphogenesis induction and shoot formation and enhanced shoot height and fresh weight. Kin had a lesser impact than BA on all parameters at all concentrations. The number of shoots differed significantly among the cultivars. The Murashige and Skoog (MS) medium supplemented with 3.0 mg/L indole-3-butyric acid (IBA) produced the highest percentage of root formation (93.3%), number of roots/plantlet (5.26), and root length (1.10 cm). The Jan Bos cultivar at 3.0 mg/L IBA had the highest mean rooting percentage (100%) and number of roots per plantlet (6.66), while Pink Pearl had the highest root length (1.39 cm).

Investigation of Growth Factors and Mathematical Modeling of Nutrient Media for the Shoots Multiplication In Vitro of Rare Plants of the Rostov Region

Abstract: Micropropagation is an effective way to preserve the gene pool of threatened plants. This study is devoted to the mathematical modeling of nutrient media and the study of the effect of mT (meta-topoline) on the multiplication of shoots of Hedysarum grandiflorum, Hyssopus cretaceus, and Matthiola fragrans in vitro in comparison with benzylaminopurine (BAP) and kinetin (KT). Initiation was performed on an MS medium with 0.5 mg/L BAP. For shoots multiplication, MS, B5, and WPM media were used with the addition of mT, BAP, KT. For H. grandiflorum, the multiplication coefficient of shoots was highest on medium B5 with the addition of mT at a concentration of 1 mg/L—2.90 shoots per plant, for H. cretaceus—B5 + 0.5 mg/L mT, and for M. fragrans—B5 + 1 mg/L KT. A positive effect of mT on H. grandiflorum and M. fragrans in vitro was found. The efficiency of using KT for H. cretaceus shoot multiplication is shown. The effectiveness of the B5 nutrient medium for H. grandiflorum and M. fragrans was determined. The positive effect of WPM for H. cretaceus micropropagation has been demonstrated. It is not recommended to use the MS media for micropropagation of these plant species.

In vitro clonal propagation of Tanacetum cinerariifolium and establishment of an ex situ collection of selected clones

Abstract: Dalmatian pyrethrum Tanacetum cinerariifolium (Trevir.) Sch. Bip. (Asteraceae) is a perennial herb endemic to the eastern coast of the Adriatic Sea. The species is widely cultivated in many countries for its bioactive compounds pyrethrins, which are used as natural insecticides. Plants derived from seeds vary greatly in pyrethrin content; therefore, the vegetative propagation of high-quality individuals is very important for the establishment of agricultural pyrethrum crops. The present study deals with rapid in vitro multiplication of pyrethrum, ex vitro adaptation of selected clones and creation of an ex situ collection, as a first step towards introducing the species into agriculture in Bulgaria. Seeds from a private ex situ collection in Bulgaria and from a natural Croatian population were used as initial material for in vitro cultures initiation. Basal MS medium (Murashige and Skoog 1962) or MS supplemented with different concentrations of kinetin and indole-3-butyric acid were used for seed germination and multiplication of one-seed derived clones by consecutive subcultivations. The propagation effectiveness was evaluated as a number of new plants obtained per initial shoot. Considerable losses were noticed due to both endophytic contaminations and necrosis, especially on media supplemented with plant growth regulators. These problems were overcome by medium optimization: adding an antibiotic and modifying the medium to increase the calcium concentration using CaCO3. In the best medium variant (basal MS + 200 mg/L Medaxone + 75 mg/L Ca) no more infected plants were observed, and the percentage of necrotic plants decreased threefold, which resulted in formation of 38.06±10.11 new plants per initial shoot for a period of 7 months. Three hundred and sixty plants were ex vitro adapted in a phytotron (88% surviving rate), then 16 plants from 4 selected clones were transferred to the ex situ collection and bloomed twice from the very first growing season (June and September). The number of the flower heads increased in the second year of field cultivation and an average of 328±138 capitula per plant were counted for the best clone. The first trials to establish a pilot plantation of pyrethrum are promising.

Micropropagation Protocol and Genetic Stability of the Salix myrtilloides Plants Cultivated In Vitro

Abstract: Simple Summary Salix myrtilloides is a relict species endangered with extinction in many Central European countries. To save the populations on the southern border of its range, in vitro propagation was used for plants production because it allows one to obtain a lot of new plants in a relatively short time without damaging the existing ones. We collected shoot pieces and multiplied them in a growing media, which contained nutrients and growth regulators. When we produced enough daughter plants, we planted them into soil and hardened them to natural conditions. Based on the conducted genetic analysis and flow cytometry, we stated that obtained plants are genetically identical to the mother ones. The conducted research confirmed that tissue culture may be used to propagate the endangered S. myrtilloides species and that the obtained plants may be used to establish new populations or to strengthen the existing ones. Abstract Salix myrtilloides L. is a relict species, threatened with extinction in many European countries. To prevent the loss of the species, tissue culture was established to produce plant material for reintroduction in natural habitats. Micropropagation was chosen as a method to obtain new plants. S. myrtilloides shoots were disinfected with NaOCl, AgNO3, or with a two-step disinfection with NaOCl, and then placed on MS medium supplemented with BA at 1 mg·dm−3 and IBA at 0.1 mg·dm−3. Regenerated shoots were cultivated in presence of BA, KIN, and 2iP to select the treatment with the highest multiplication rate. The obtained plants were acclimatized to ex vitro conditions. Inter-simple sequence repeat (ISSR) and flow cytometric analyses were conducted on in vitro regenerated plants to check their genetic stability. The best disinfection results were obtained when explants were treated with 1.5% NaOCl for 20 min. The highest multiplication rate and good quality plants were noted in the control media, without growth regualtors and in presence of kinetin at 0.5 mg·dm−3. Flow cytometry and ISSR analyses confirmed genetic stability in plantlets, which indicated the possibility to use the in vitro obtained plants for reintroduction.

Induction of Axillary Bud Swelling of Hevea brasiliensis to Regenerate Plants through Somatic Embryogenesis and Analysis of Genetic Stability

Abstract: To overcome rubber tree (RT) tissue culture explant source limitations, the current study aimed to establish a new Hevea brasiliensis somatic embryogenesis (SE) system, laying the technical foundation for the establishment of an axillary-bud-based seedling regeneration system. In this study, in vitro plantlets of Hevea brasiliensis Chinese Academy of Tropical Agricultural Sciences 917 (CATAS 917) were used as the experimental materials. Firstly, the optimum conditions for axillary bud swelling were studied; then, the effects of phenology, the swelling time of axillary buds (ABs), and medium of embryogenic callus induction were studied. Plantlets were obtained through somatic embryogenesis. Flow cytometry, inter-simple sequence repeat (ISSR molecular marker) and chromosome karyotype analysis were used to study the genetic stability of regenerated plants along with budding seedlings (BSs) and secondary somatic embryo seedlings (SSESs) as the control. The results show that the rubber tree’s phenology period was mature, and the axillary bud induction rate was the highest in the 2 mg/L 6-benzyladenine (6-BA) medium (up to 85.83%). Later, 3-day-old swelling axillary buds were used as explants for callogenesis and somatic embryogenesis. The callus induction rate was optimum in MH (Medium in Hevea) + 1.5 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) + 1.5 mg/L 1-naphthalene acetic acid (NAA) + 1.5 mg/L Kinetin (KT) + 70 g/L sucrose (56.55%). The regenerated plants were obtained after the 175-day culture of explants through callus induction, embryogenic callus induction, somatic embryo development, and plant regeneration. Compared with the secondary somatic embryo seedling control, axillary bud regeneration plants (ABRPs) were normal diploid plants at the cellular and molecular level, with a variation rate of 7.74%.

In Vitro Seed and Clonal Propagation of the Mediterranean Bee Friendly Plant Anthyllis hermanniae L.

Abstract: Anthyllis hermanniae L. (Fabaceae) is a perennial Mediterranean shrub with the potential to be used as a bee-friendly ornamental plant in arid and semi-arid regions, valued for its tolerance of barren soils, winds, and strong temperature changes. With the aim of facilitating the introduction of the species into the horticulture industry, its in vitro seed and clonal propagation was investigated for the first time, to our knowledge. Seeds stored in the dark at room temperature for 4, 7, 9, 12, and 18 months germinated at percentages higher than 80% after scarification, when incubated in vitro in solid half-strength Murashige and Skoog (MS) medium at temperatures from 10 to 25 °C, while photoperiod (continuous darkness or 16 h light period/8 h dark) during incubation did not affect germination. Explants excised from in vitro grown seedlings established at higher percentages compared to explants from adult native plants, more efficiently in MS medium with 1.0 mg L−1 6 N benzyladenine (BA). During subcultures in the same medium, juvenile explants formed more and longer shoots than adult ones. Almost all adult explants formed shoots when subcultured in MS medium with 0.0 to 4.0 mg L−1 BA, zeatin, kinetin or 6-(γ,γ-dimethylallylamino)purine (2iP). ΒΑ at 0.5 to 2.0 mg L−1 induced many more shoots (17–21) per explant and much higher multiplication indices compared to all other cytokinins, while longer shoots were produced in a medium without hormones or with 0.5–1.0 mg L−1 2iP. Microshoots cultured in half-strength MS medium with 0.0–4.0 mg L−1 indole-3-butyric acid rooted at highest percentage (around 70%) in the medium containing 4.0 mg L−1 IBA, while microshoots of juvenile origin developed more and longer roots compared to adult ones. Micropropagated plantlets were successfully acclimatized ex vitro (>97%), regardless of their origin. The efficient micropropagation of A. hermaniae will facilitate its sustainable exploitation as a bee-friendly landscape plant, a forage plant for honeybees in Mediterranean areas, and a medicinal plant.

Effect of Plant Growth Regulators on Cherry Tomato (Solanum lycopersicum var. cerasiforme)

Abstract: A cherry tomato is a small-sized tomato variety known for its round shape and sweet flavor. Therefore, at the Department of Horticulture at Naini Agricultural Institute, Sam Higginbottom University of Agriculture Technology and Sciences, Prayagraj, Uttar Pradesh, an investigation titled "Effect of Plant Growth Regulators on Cherry Tomato" was conducted during the Rabi-2022-23 season to assess the impact of plant growth regulators on the growth, quality, and yield of Pusa Cherry 1 tomatoes, as well as analyze the economic aspects of different treatments. Ten treatments were employed, including control with different concentrations of growth regulators (GA3 and Auxin) and Kinetin. The study found that using plant growth regulators had a significantly positive effect on cherry tomatoes for all traits. Among the treatments tested, T3(GA3 @ 75 ppm) showed the most favourable results in terms of plant height, number of primary branches, early flowering and maturing. T3 also demonstrated superior performance in terms of fruit weight, the maximum number of fruits per plant, and yield per plant.

Kinetin Capped Zinc Oxide Nanoparticles Improve Plant Growth and Ameliorate Resistivity to Polyethylene Glycol (PEG)-Induced Drought Stress in Vigna radiata (L.) R. Wilczek (Mung Bean)

Abstract: Plants are sessile and mostly exposed to various environmental stresses which hamper plant growth, development, and significantly decline its production. Drought stress is considered to be one of the most significant limiting factors for crop plants, notably in arid and semi-arid parts the world. Therefore, the present study aimed to evaluate the potential impact of different concentrations (10, 100, and 200 µg/mL) of kinetin capped zinc oxide nanoparticles (Kn-ZnONPs) on Vigna radiata (L.) R. Wilczek under varying levels (5%, 10%, 15%) of PEG-induced drought stress. ZnONPs were synthesized by a co-precipitation method using Zinc acetate as a precursor at pH-12, incinerated to 500 °C, and kinetin was used as a surface functionalizing agent. The resulting Kn-ZnONPs were characterized by various contemporary analytical techniques, including SEM, SEM-EDS, XRD, DLS, and Zeta potential and IR spectroscopy. Crystalline Kn-ZnONPs, with a zeta potential of 27.8 mV and a size of 67.78 nm, of hexagonal wurtzite structure and vibrational stretches associated with N-H, C-O, C-N, etc., were confirmed. PEG-induced drought stress significantly reduced the growth of V. radiata by declining the chlorophyll and carotenoid contents. Moreover, a significant decrease in the levels of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), soluble sugar contents, proline, protein contents, phenol, and tannin were observed compared to the control. However, the exogenous application of Kn-ZnONPs ameliorated all photosynthetic parameters by up-regulating the antioxidant defense system through the promotion of SOD, POD, CAT, and lipid peroxidation levels. The biochemical parameters, such as proteins, soluble sugars, and proline, were observed to be maximum in plants treated with 200 µg/mL Kn-ZnONPs under 5% drought stress. The application of Kn-ZnONPs also enhanced the total phenol contents, flavonoid, and tannin contents. In conclusion, the findings of this study demonstrate that the exogenous application of Kn-ZnONPs provides beneficial effects to V. radiata by attenuating the damaging effects of drought stress through the up-regulation of the antioxidant defense system and osmolytes. These results suggest that Kn-ZnONPs have potential as a novel approach to improve crop productivity under drought stress conditions.

In vitro development of gametophytes and sporophytes of critically endangered fern Christella kendujharensis S.K. Behera & S.K. Barik (Thelypteridaceae)

Abstract: Christella kendujharensis S.K. Behera & S.K. Barik (family: Thelypteridaceae) is a critically endangered fern endemic to India. We developed its mass propagation protocol through in vitro spore culture in a liquid suspended medium for germination of gametophytes that produced sporophytes subsequently. The spores were monolete, reniform (mean length 36.49 ± 3.6 × breadth 25.67 ± 4.2 μm) to spheroidal with fimbriate and broad-winged perispore. The maximum germination was in the Parker's and Thompson's (P&TV) medium with 0.1 mg L−1 thiamin, 0.1 mg L−1 riboflavin, 0.1 mg L−1 biotin, 0.1 mg L−1 pyridoxin, 0.5 mg L−1 nicotinic acid, 100 mg L−1 myoinositol, 0.01 mg L−1 folic acid, and 3 mg L−1 glycine. Spore germination was Vittaria type and gametophyte development was Aspidium type. The cordate gametophyte developed archegonia on the 38th day and antheridia on the 42nd day of spore sowing. Both isolate and composite cultures developed sporophytes indicating the presence of intra-gametophytic selfing. Based on the maximum number of regenerated gametophytes, two combinations, that is, (i) 2 mg L−1 kinetin, and 0.01 mg L−1 2,4-dichlorophenoxyacetic acid, and (ii) 0.1 mg L−1 naphthaleneacetic acid and 2 mg L−1 kinetin in P&TV media were recommended for mass propagation of C. kendujharensis.

A study on the bud regeneration ability of Cymbidium wenshanense by in vitro culture

Abstract: The study on the buds regeneration of Cymbidium wenshanenseorchid seedlings cultured in vitro in the conditions of 25oC room temperature, 65% humidity, 2000 lux light intensity, and 16 hours of lighting a day was implemented with the aim of finding out the proper medium, cytokinin growth stimulators (kinetin, TDZ, and BA), and sucrose concentrations on seed regeneration. Results showed that MS medium had a good effect on seed germination (the highest is 90.54%) and cytokinin BA significantly improved the buds regeneration of Cymbidium wenshanense orchid (4.42 buds of 3.63 cm length, and 272.67 mg weight formed from a sample when 1 mg of BA was added). In addition, sucrose supplemented at 5% concentration was considered to be successful (4.47 buds of 3.6 cm length and 315 mg were produced when 5% sucrose was added). This is a result of practical significance in the conservation and development of Cymbidium wenshanenseorchid in north Vietnam.

Shoots and Roots Induction of Garlic on Different Composition of Plant Growth Regulators and Photoperiod

Abstract: Plant Growth Regulators and photoperiods will affect callus morphogenesis from garlic bulbils. The study aimed to obtain the best interaction between media composition and photoperiod for shoots and root induction of the garlic callus. The research has two-stage. The first stage consisted of two factors. The first factor is the combination of PGR for shoots induction which consists of three combinations, i.e., 0.1 ppm NAA + 1 ppm BAP, 2 ppm NAA + 2 ppm BAP, and 0.4 ppm 2,4-D + 2 ppm kinetin. The second factor is photoperiod which consists of 10, 16, and 22-h light conditions. The second stage is the root induction in different PGRs (2 ppm IBA and 0.2 ppm NAA + 2.25 ppm BA) under continuous dark conditions. The results of the first stage showed that the PGR produced the most shoots was 0.1 ppm NAA + 1 ppm BAP. After four weeks, the frequency of an explant emerging during the shoot reached 64% on media 0.1 ppm NAA + 1 ppm BAP. The treatment MS media supplemented with 0.1 ppm NAA + 1 ppm BAP at 16-h photoperiods increased the growth of shoots per explant in 11-15 shoots and produced the most extended shoot length. The result of the second stage showed that the number of roots on 2 ppm IBA is the same as NAA 0,2 ppm + BA 2,25 ppm, but the treatment 2 ppm IBA has the highest root number in three months.

Effectiveness of Soaking with Gibberellic Acid and Kinetin on Germination and Growth Indicators of Two Cultivars of Pistachio Plant Pistaciavera L.

Abstract: The study was conducted in the plant canopy of the Department of Horticulture and Landscape/College of Agriculture/University of Kerbala during the season 2021-2022 to study the effect of three factors, the first factor is two cultivars (Ajami and Nab Al-Jamal) and the second factor is gibberellic acid in three concentrations (0,100,200 mg. L-1) respectively, and the third factor is kinetin in three concentrations and symbolizes (0, 15, 30 mg. L-1) respectively. The results of the statistical analysis showed the following: Nab Al-Jamal cultivar, kinetin at a concentration of (15) mg. L-1 and gibberellin at concentration (100 and 200) mg. L-1 were superior in most of the studied traits as single factors. For bilateral interactions, the concentration of gibberellin with kinetin is (200 + 15) mg. L-1 has outperformed in most of the studied traits. As for the interaction of the cultivar with gibberellin, the interaction treatment (Nab Al-Jamal + 100 mg. L-1 gibberellin) gave the best results in the percentage of germination, speed of germination and germination energy, while the treatment (Ajami + 200 mg. L-1 gibberellin) gave the best results in plant height and stem diameter, while the interaction treatment (Nab Al-Jamal +200 mg. L-1 gibberellin) gave the best results in the number of leaves. The triple interaction was (Ajami + 200 gibberellin + 15 kinetin) mg. L-1 was superior in the results of the following traits: percentage of germination, germination energy, stem diameter and number of leaves. As for the overlap treatment (Nab Al-Jamal + 100 gibberellin + 0 kinetin) mg. L-1 has outperformed in germination speed and plant height.

Micropropagation and assessment of genetic stability of Dendrobium transparens Wall. Ex Lindl. using RAPD and ISSR markers

Abstract: Introduction Dendrobium species have been widely used for many health disorders since ancient times. However, due to unrelenting collection to meet the increasing demand for their use in medication and other health products, the natural habitats of medicinal Dendrobium transparens have been devastated and are on the verge of extinction. Methods An efficient in-vitro propagation protocol for Dendrobium transparens using seed derived protocorms was established and genetic homogeneity of the in-vitro regenerants and the wild plant was studied. Results The maximum seed germination was observed in Full strength Murashige and Skoog medium (FMS). Induction of protocorms were achieved on basal as well as half-strength MS medium. The highest number of shoot (11.9 shoots/explant) was achieved in half MS medium fortified with 100 mL/L coconut water in addition with Benzyl amino purine (BAP) 1 mg/L and Kinetin 2 mg/L. Further, elongated shoots were transferred to full and half strength MS root initiating medium supplemented with different concentration of auxins. However, a maximum of (8.3 ± 0.6, 4.9 ± 0.1 cm) roots were achieved in full MS medium fortified with 100 mL/L coconut water and Napthalene acetic acid (NAA) 1.5 mg/L. Ten rapid Random Amplified Polymorphic DNA (RAPD) and Inter Simple Sequence Repeats (ISSR) primers were used to analyze genetic stability among in-vitro and mother plant. RAPD primers produced a total of 23 fragments while ISSR primers produced a total of 16 fragments. Conclusion The amplified bands of all the samples of in-vitro plants were similar to bands of mother plant. The present research reported here is indicating the applicability of tissue culture for true-to-type plant production and conservation of D. transperens.

Secondary Metabolite Production in Callus Culture of Verbascum scamandri Murb.

Abstract: Verbascum scamandri Murb. known as "Kazdağı Mullein" is an endemic species in Türkiye and is classified as an endangered (EN) species. The aim of this study is to establish an efficient callus culture for V. scamandri and analyze the amounts of verbascoside, luteolin, and aucubin metabolites of calli samples. Leaf explants were cultured on MS medium with cytokinin (BAP, Kin, 0, 0.5, 1, 2, 3 mg/L) and auxin (NAA, 2,4-D, 0, 0.1, 0.5, 1 mg/L), 1 g/L PVP, 3% sucrose, and 0.7% agar for callus induction. Callus tissue in MS with 2 mg/L Kin, 0.5 mg/L Kin+0.5 mg/L 2,4-D, 2 mg/L Kin+0.5 mg/L 2,4-D, and 3 mg/L Kin+0.5 mg/L 2,4-D was proliferated in MS basal medium containing PGR at the same concentrations and combinations as the callus induction media. Verbascoside, luteolin, and aucubin were quantified in leaf samples of the in vivo collected plants, leaf samples of in vitro growing plants, and calli using HPLC-DAD. According to the results, the verbascoside content in the leaf of collected plants was 7.03 mg/g, luteolin was 0.66 mg/g, and aucubin was 2.99 mg/g. The leaf of in vitro plants had 1.62 mg/g verbascoside, 0.18 mg/g luteolin, and 1.32 mg/g aucubin. Whereas, the maximum content of secondary metabolites in the callus samples was observed 13.77 mg/g verbascoside in MS medium with 2 mg/L Kin, 0.51 mg/g luteolin in MS medium with 2 mg/L Kin+0.5 mg/L 2,4-D, and 9.32 mg/g aucubin in 0.5 mg/L Kin+0.5 mg/L 2,4-D.

In vitro propagation and genetic fidelity analysis of Celastrus paniculatus Willd – a vulnerable high trade medicinal woody liana of Western Ghats

Abstract: Celastrus paniculatus Willd, a vulnerable important woody liana with high medicinal value and limited reproductive capacity has been over exploited due to its demand in the pharmaceutical industry. In the present study, an efficient micropropagation protocol was developed from nodal explants on Murashige and Skoog and woody plant medium. The effect of various strengths of WP and MS medium (total, half strength) and different concentrations of plant growth regulators including cytokinins [6-benzylaminopurine (BAP) and Kinetin) and auxins (α-naphthaleneacetic acid (NAA) and (IBA) indole-3-butyric acid] on proliferation and rooting was investigated. C. paniculatus shoots showed maximum proliferation in WP medium supplemented with BAP (0.5 mg l-1). Best rooting frequency and higher root number and length were attained in medium fortified with NAA in combination with 0.2% activated charcoal. Seventy percent of the plantlets were successfully acclimatized to ex vitro conditions, exhibiting normal development. These plantlets can be used to replenish the declining populations in the wild. Molecular profiling of the in vitro developed plantlets was done using RAPD to confirm their genetic uniformity.

Establishment of an in vitro method for micropropagation of ironwort, (Sideritis raeseri Boiss. & Heldr.)

Abstract: Ironwort / Mountain Tea (Sideritis raeseri Boiss & Heldr.) is an endangered (EN) plant species in Albania. This study aimed to develop a rapid clonal propagation protocol using in vitro methodologies. The ironwort seeds were pre-treated with three concentrations of GA3 (250, 500, and 1000 mg l-1). During the inoculation stage, two types of culture media, Murashige & Skoog (MS) and Woody Plant Medium (WPM), were tested, and the effects of both GA3 concentration and culture media used were evaluated. For the subculture stage, three cytokinins (6-benzylaminopurine / BAP, kinetin, zeatin) at four concentrations (0.5; 1.0; 1.5; 2.0 mg l-1), were compared for the RGR index, while for the rooting stage, two different auxins (1-naphthaleneacetic acid / NAA and indole-3-butyric acid / IBA) at four concentrations (0.5; 1.0; 1.5; 2.0 mg l-1) were tested. GA3 at 500 mg l-1 and MS medium resulted as more effective. The highest value of the RGR index during the subculture stage was obtained in the MS nutrient medium supplemented with BAP at 1.5 mg l-1. For rhizogenesis response, IBA was more effective for roots and length number. Based on these results, in vitro methodologies can be a promising tool for the mass production of this endangered plant species and with possible applications for enhancing the production of valuable nutraceuticals.

In Vitro Propagation and Genetic Uniformity Assessment of Manglietiastrum sinicum: A Critically Endangered Magnoliaceae Species

Abstract: Manglietiastrum sinicum Y.W. Law is a critically endangered species with great ornamental and commercial value, which urgently requires protection. We tested different combinations of basal media and plant growth regulators to determine (i) the optimal conditions for bud induction and proliferation of explants and (ii) optimal rooting conditions. RAPD- and ISSR-PCR were used to assess the genetic fidelity of regenerated plantlets. Murashige and Skoog medium (MS) supplemented with 0.5 mg/L 6-benzyladenine (BA) and 0.05 mg/L indole-3-butyric acid (IBA) is the optimal medium for bud induction (100% induction). MSM medium (a special basal medium for M. sinicum) was more suitable for the efficient proliferation and rooting of M. sinicum. Maximum bud proliferation rate (446.20%) was obtained on MSM, with 0.4 mg/L BA, 0.5 mg/L kinetin, and 0.06 mg/L IBA, while maximum root induction rate (88.89%) was obtained on MSM supplemented with 0.4 mg/L 1-naphthylacetic acid and 1.0 mg/L IBA with a 7-day initial darkness treatment. The rooted plantlets were transferred to a substrate containing peat soil, perlite, coconut chaff, and bark (volume ratio 2:1:1:1), with a resulting survival rate of 92.2%. RAPD and ISSR markers confirmed the genetic uniformity and stability of regenerated plants.