Search Results (70)

Grapevine breeding programs face difficulties in preserving germplasm, especially from species and interspecific hybrids, since most collections are maintained in the field and exposed to biotic and abiotic stress, which can lead to material loss. The Universidade Estadual do Norte Fluminense Darcy Ribeiro (UENF) Grapevine Breeding Program faces similar challenges, limiting studies on hybrids resistant to the nematode Pratylenchus brachyurus and downy mildew (Plasmopara viticola), which are valuable for genetic improvement. This study aimed to implement in vitro conservation under minimal growth conditions for interspecific hybrids of Vitis spp. from the UENF program. The protocol followed a completely randomized design in a 2 × 2 × 3 factorial scheme: two hybrids (CH1.2 and CH1.3), two temperatures (18 ± 1 °C and 27 ± 2 °C), and three sucrose concentrations (10, 20, and 30 g L⁻¹), over 180 days of in vitro culture. The results showed that conservation of the UENF hybrids is feasible using nodal segments as explants, at 18 ± 2 °C and 10 g L⁻¹ of sucrose, for up to four months. This protocol may also be applied to other Vitis spp., contributing to the preservation and continued study of valuable germplasm. Full article

The introduction of new ornamental species and cultivars is one of the hallmarks of innovation in global floriculture. Brunfelsia uniflora, a subshrub native to Brazil, has white, lilac, and blue flowers on the same plant, in addition to a distinctive fragrance. As it is a wild species, technologies such as large-scale clonal propagation of superior genotypes are still scarce, limiting its supply to the flower market. Therefore, a successful micropropagation protocol was developed for B. uniflora using nodal segments and shoot tips as initial explants. In the multiplication phase, the use of 6-benzylaminopurine produced the highest multiplication rates (10.3–10.9 shoots/explant) and the number of leaves in the shoots. In vitro shoot rooting using MS medium with reduced macronutrient concentrations and supplemented with IBA resulted in a 91.7% rooting rate. The greatest difficulty in micropropagating this species was the high percentage of shoots that developed calli. The highest percentage of callus formation occurred with the addition of auxins at high concentrations (1.0 and 1.5 mg L⁻¹). Even so, the shoots and plantlets were acclimatized, demonstrating the effectiveness of this technique for the production of B. uniflora plantlets. Full article

Triadica sebifera, an economically and medicinally valuable tree species native to China, was investigated for its in vitro regeneration potential using leaf explants from nodal cuttings of young stems and sprouts. This study evaluated the effects of basal media, plant growth regulators (PGRs), explant sources, and incision methods on adventitious shoot induction, supplemented by histological analysis. The highest shoot regeneration frequency (98.89%) and maximum shoot number (72) were achieved via direct organogenesis using sucker-derived nodal cuttings cultured on MS medium with 2 mg/L 6- benzyladenine (6-BA), 0.3 mg/L kinetin (KT), and 0.2 mg/L α-naphthaleneacetic acid (NAA). Under identical conditions, branch-derived explants showed lower regeneration (84.44%, 64 shoots). Transverse midvein incision proved most effective, with sucker-derived leaves exhibiting superior regeneration. Shoots elongated completely (100%) on Murashige and Skoog (MS) medium containing 0.3 mg/L 6-BA, 0.03 mg/L NAA, and activated charcoal. Rooting was optimal on MS medium with 0.3 mg/L indole-3-butyric acid (IBA), yielding a 98% acclimatization survival rate. Histological analysis revealed de novo meristem formation from parenchyma cells, confirming direct organogenesis without callus intermediation, further validating the enhanced regenerative capacity of sprout-derived explants. This efficient in vitro regeneration system provides a foundation for large-scale propagation and germplasm conservation of T. sebifera, while offering insights for woody plant regeneration studies. Full article

Hyperhydricity is characterized by morphological abnormalities and reduced plant vigour. This study investigated the use of a bottom cooling system (creating an approximate 2 °C temperature differential) during culture initiation to evaluate the impact on hyperhydricity in cannabis micropropagation. Nodal explants from two clonal triploid cultivars known to exhibit hyperhydricity, Higher Education 1 (HED-1) and Higher Education 2 (HED-2), were surface sterilized and placed in culture tubes using standard methods. Treatments included bottom cooling, metal pads without bottom cooling, and standard shelving (controls—no pad). Various morphological and physiological traits were assessed, including a detached leave water loss assay, dry mass, chlorophyll content, and survival rate. Plants cultured with bottom cooling showed significantly higher survival rates, healthier appearance, and improved physiological parameters compared to controls. In contrast, many control explants were hyperhydric with translucent and brittle leaves. Quantitative data revealed significant improvements in fresh weight (54.84% for HED-1 and 51.42% for HED-2), dry weight (36% for HED-1 and 8% for HED-2), chlorophyll fluorescence ratios (7.24% for HED-1 and 9.18% for HED-2), chlorophyll content (18.38% for HED-1 and 20.67% for HED-2), and cuticle/stomate function (30% for HED-1 and 27.27% for HED-2) using bottom cooling. Moreover, our morphological observation showed that almost 85% of control plants were hyperhydric, whereas only 10% of the plants cultured with a bottom cooling system were hyperhydric. This study confirmed that bottom cooling helps reduce the rate and impacts of hyperhydricity in cannabis and significantly improves the survival and quality of in vitro plants. Full article

“GiSelA 17” (Prunus canescens × Prunus avium) is a novel cherry clonal rootstock with the ability to bear fruit early and resist replant situations, and it has a high tolerance to the menaces of Prunus dwarf virus (PDV) and Prunus necrotic ring spot virus (PNRSV). In this study, two kinds of explants were taken, i.e., shoot tip (E1) (10 mm) and nodal segment (E2) (15 mm) explants. Five different sterilant regimes using sodium hypochlorite, mercuric chloride, and ethyl alcohol were employed to assess surface sterilization. Two types of media, namely Murashige and Skoog (MS) and Woody Plant Medium (WPM), and twelve and six plant growth regulator combinations with benzyl amino purine (BAP) and indole-3-butyric acid (IBA) were used, respectively, for the establishment and proliferation steps. The results show that maximum culture asepsis (75.33%) was obtained with shoot tips (E1) using 0.05% HgCl₂ for 5 min + 70% ethanol for 10 s (S4), and maximum explant survival (80.33%) was observed in 0.1% HgCl₂ for 5 min (S1) for shoot tips (E1). The maximum establishment rate (83.33%) was found in shoot tips (E1) in MS medium with BAP + IBA (1 + 0.01 mg/L) during the establishment step, with a maximum proliferation rate of 92.00% obtained in MS and BAP (0.75 mg/L). Inferior establishment results (26.66%) were obtained in nodal segments (E2) using WPM and BAP + IBA (1.50 + 0.01 mg/L), with a low proliferation rate (68.66%) in WPM and BAP + IBA (0.25 + 0.01 mg/L). Nonetheless, our research is the first in vitro study on “GiSelA 17” rootstock that focuses on generating the best quality planting material for commercial cherry production. Full article

Potatoes, a vital global food crop, have shown remarkable adaptability, significantly contributing to food security. Technological advancements now enable their cultivation from soil-based systems to liquid synthetic nutrient media, even in artificial closed environments without natural light or fertile soil. This study examined the effects of Benzylaminopurine (BAP) and Kinetin (Kin) at concentrations ranging from 0 to 5 mg/L and sucrose concentrations ranging from 20 to 120 g/L on in vitro tuberization, focusing on microtuber size, weight, and tuberization rate. Nodal segments from virus-free ‘Red Scarlet’ in vitro potato plantlets were used as explants. These explants were cultured on Murashige and Skoog (MS) medium solidified with 0.5% agar. The study also compared minituber production efficiency under soil-based greenhouse and aeroponic conditions. The highest in vitro potato tuberization rate (90%) was achieved with 80 g/L sucrose and 3.0 mg/L BAP. After induction, virus-free microtubers were transferred to both greenhouse conditions and aeroponic systems for further assessment of minituber production and biochemical composition. These findings demonstrate the potential of aeroponics as a superior method for producing high-quality, pathogen-free minitubers. Aeroponics resulted in significantly higher minituber yields compared to soil-based greenhouse systems, offering a scalable and efficient solution for seed production. Full article

In recent years, interest in Lonicera caerulea production has grown significantly because of its nutritional and pharmaceutical benefits, leading to rapid expansion in its cultivation. L caerulea var. emphyllocalyx is a lesser-known botanical variety. Due to differences between plants of the Lonicera genus and the lack of scientific reports on micropropagation, it is necessary to determine the possibilities of in vitro propagation. The aim of this study was to elaborate a micropropagation protocol of two new breeding clones of Lonicera caerulea var. emphyllocalyx: ‘21–17’ and ‘139–24’. The experiments were carried out on in vitro cultures grown on MS medium supplemented with 1 mg·dm⁻³ BA or 1 mg·dm⁻³ mT. Two types of explants were used during the experiment: nodal fragments (NFs) and shoot-tips (STs). Before acclimatisation, some rooted microshoots were subjected to cooling at 4 °C for 4 weeks. Significantly more ST explants than NF explants started to grow at the proliferation stage. The application of BA resulted in much better proliferation and health of cultures. Cold storage of micropropagated ‘139–24’ plantlets significantly increased their survival in acclimatisation in contrast to ‘21–17’ plantlets but weakened further growth of the plants. In future in vitro studies on L. caerulea var. emphyllocalyx, BA can be used as the primary growth regulator due to its effectiveness and low cost. Nodal fragments should be considered as the main propagation material since they promote better growth rates. Additionally, further research is required to explore the effects of low-temperature storage on the growth and physiology of these plants. The results obtained in this research may contribute to the development of micropropagation technology in the future for L. caerulea var. emphyllocalyx. Full article

Dianthus cruentus Griseb. (Caryophyllaceae) is an herbaceous perennial native to Greece with a strong ornamental potential when used as a pollinator-friendly component of xeric gardens and green roofs, where it is valued for its tolerance of poor, dry soils, and its showy colorful inflorescences. Aiming to develop an efficient mass propagation protocol appropriate for the introduction of the species as a novel floricultural crop, the in vitro seed and clonal propagation of a Greek native xeric ecotype were investigated in this paper for the first time. A total of 90–100% of the seeds, after being stored in the dark at room temperature for 12 months, germinated when incubated at 10 to 25 °C after their surface sterilization and transfer in vitro. Sixty-day-old seedlings grown in vitro were then used as a source of nodal explants for the initial establishment of micropropagation cultures, more efficiently on MS medium with 0.1 mg L⁻¹ 6-benzylaminopurine (BA). In the multiplication stage, either normal or hyperhydric micro-shoots were used as explant sources, assessing the possibility of incorporating usually discarded material in the propagation procedure. Different solid media were tested, with the highest multiplication indices (5.1) recorded in an MS medium containing 0.1 mg L⁻¹ BA and 0.05 mg L⁻¹ NAA, regardless of explants’ hyperhydricity, while an MS medium containing 0.1 mg L⁻¹ BA and 12 g L⁻¹ agar proved optimal for the effective reversal of hyperhydric explants (MI: 5.2). Despite higher hyperhydricity and reaction rates being observed when hyperhydric explants were used, modifications in the multiplication medium proved to be highly effective in controlling hyperhydricity, with the highest number of normal shoots (2.4–2.6) produced in BA-containing media. Micro-shoots rooted readily in ½ MS medium (60–100%), with rooting rates and quality positively affected by the presence of 0.5 mg L⁻¹ IBA in the rooting medium and the absence of cytokinins in the multiplication one. Rooted micro-shoots were successfully acclimatized ex vitro at high rates (65–100%), their origin influencing their acclimatization and morphology. Thus, the concurrent use of normal and hyperhydric shoots in the proposed micropropagation protocol is proven to be both feasible and desirable, as it is able to significantly increase efficiency and facilitate the sustainable exploitation and dissemination of D. cruentus as a promising multivalent horticultural crop. Full article

Aseptic seedlings of different ages derived from surface-sterilized mature seeds were applied as an explant source. Various explants such as 7- and 21-day-old hypocotyl fragments, 42-day-old nodal stem segments, and transverse nodal segments of stem, as well as leaf petioles, were cultured on the agar-solidified Murashige and Skoog (MS) basal medium supplemented with 0.1 mg/L IAA, 5 mg/L AgNO₃ and different types and concentrations of cytokinin (1 mg/L zeatin, 0.25 mg/L thidiazuron (TDZ), and 5 mg/L 6-benzylaminopurine (6-BAP)). Consequently, it was found that 7- and 21-day-old hypocotyl fragments, as well as nodal stem segments obtained from adult aseptic seedlings, are characterized by a high explant viability and callus formation capacity with a frequency of 79.7–100%. However, the success of in vitro somatic shoot organogenesis was significantly determined not only by the culture medium composition and explant type but also depending on its age, as well as on the size and explant preparation in cases of hypocotyl and age-matched nodal stem fragments, respectively. Multiple somatic shoot organogenesis (5.7 regenerants per explant) with a frequency of 67.5% was achieved during 3 subcultures of juvenile hypocotyl-derived callus tissue on MS culture medium containing 0.25 mg/L TDZ as cytokinin source. Castor bean regenerants were excised from the callus and successfully rooted on ½ MS basal medium without exogenous auxin (81%). In vitro plantlets with well-developed roots were adapted to ex vitro conditions with a frequency of 90%. Full article

The efficiency and method of regeneration in in vitro culture conditions depend primarily on the plant growth regulators (PGRs) used. Even growth regulators belonging to one group may have different effects, stimulating the process of direct or indirect organogenesis, thus possibly disturbing the genetic stability among regenerants. The main aim of this study was to identify the genetic stability of Scutellaria baicalensis regenerates obtained by in vitro culture method using start codon targeted (ScoT) markers. S. baicalensis nodal explants were regenerated on MS medium supplemented with kinetin (KIN) at concentrations of 0.25, 0.5, 1.0, and 2.0 mg × dm⁻³ or benzylaminopurine (BAP)—0.25, 0.5, 1.0, and 2.0 mg × dm⁻³. The effects of the number of propagated shoots, length, number of nodes, and fresh mass of regenerants were assessed. Moreover, the genetic stability of the regenerants was analyzed using start codon targeted (SCoT) markers. Direct shoot organogenesis was observed on an MS medium containing kinetin, while indirect shoot induction occurred on an MS medium supplemented with BAP. The highest average number of shoots (3.6) was achieved for the MS + KIN medium at a concentration of 0.25 and 5.8 for the MS + BAP 1.0 medium. The average length and average number of nodes were the highest on the MS + BAP 0.25 medium (50.0 and 6.0, respectively), while the lowest values of these features were observed on the MS + KIN 2.0 medium (40.3 and 4.9, respectively). A total of 111 amplified bands were exhibited by SCoT primers. Three of the analyzed primers revealed four unique genotype-specific markers. The average percentage of polymorphism obtained was 36.7%. The analysis of genetic similarity revealed a high level of genetic similarity between the donor plant and regenerants obtained on MS “0” (medium without the addition of phytohormones). A slightly lower value of genetic similarity was observed for regenerants obtained by direct organogenesis (MS + KIN medium at all concentrations). Indirect shoot organogenesis observed on the MS + BAP medium (all concentrations) resulted in the highest differentiation, both in relation to the donor plant and MS “0” regenerants. The results of our work indicate that, in the case of S. baicalensis, the maintenance of genetic stability depends primarily on the presence of the cytokinin type in the medium. Full article

Plumbago indica L. contains a valuable bioactive compound called plumbagin. Elicited regenerated shoots grown in vitro could be another source of high-yielding plumbagin. The purpose of this investigation was to examine the effects of elicitor type and concentration, as well as elicitation period, on plumbagin content in in vitro-regenerated shoots of P. indica. Nodal explants were cultured on Murashige and Skoog (MS) medium containing 1 mg/L benzyladenine (BA) in combination with 0–150 mg/L yeast extract or 50–150 µM salicylic acid for four weeks. Plumbagin levels of 3.88 ± 0.38% and 3.81 ± 0.37% w/w g dry extract were achieved from the 50 and 100 mg/L yeast extract-elicited shoots, which were higher than the value obtained for the control. However, the addition of salicylic acid did not increase the plumbagin content. In the elicitation period experiment, nodal explants were cultured on MS medium supplemented with 1 mg/L BA and 50 mg/L yeast extract for durations of three, four and five weeks. The 4-week yeast extract-elicited shoot had a maximum plumbagin content of 3.22 ± 0.12% w/w g dry extract, greater than that of the control. In summary, the plumbagin content of the in vitro P. indica shoots was enhanced by 4-week elicitation using 50 mg/L yeast extract. Full article

The damask rose (Rosa damascena Mill.) is an ornamental–medicinal plant from the Rosaceae family, and its aromatic compounds and essential oils are applied globally in the food, cosmetic, and pharmaceutical industries. Due to its economic value, this research aimed to establish a protocol for an efficient, rapid, and cost-effective method for in vitro shoot multiplication and rooting of the R. damascena ‘Kashan’ and ‘Hervy Azerbaijan’ genotypes. Nodal segments (as primary explants) were cultured on the Murashige and Skoog (MS) medium with combinations of various plant growth regulators (PGRs) such as gibberellic acid (GA₃), 6-benzylaminopurine (BAP), and indole-3-butyric acid (IBA), as well as a PGR-like substance, phloroglucinol (PG), vitamins such as ascorbic acid (AA), and activated carbon in the form of active charcoal (AC). For the establishment stage, 0.1 mg·L⁻¹ PG, 0.2 mg·L⁻¹ GA₃, and 1 mg·L⁻¹ BAP were added to the media. Secondary explants (nodal segments containing axillary buds produced from primary explants) were obtained after 30 days of in vitro culture and transferred to the proliferation media supplemented with different concentrations of BAP (0, 0.5, 1, 1.5, 2, and 2.5 mg·L⁻¹) and GA₃ (0, 0.1, 0.2, 0.4, 0.8, and 1 mg·L⁻¹) together with 0.1 mg·L⁻¹ PG and 20 mg·L⁻¹ of AA. The rooting media were augmented with different concentrations of BAP and GA₃ with 0.1 mg·L⁻¹ of IBA, PG and 20 mg·L⁻¹ of AA and AC. The results showed that the highest regeneration coefficient (4.29 and 4.28) and the largest number of leaves (23.33–24.33) were obtained in the explants grown on the medium supplemented with 2 mg·L⁻¹ BAP and 0.4 mg·L⁻¹ GA₃ for the ‘Kashan’ and ‘Hervy Azerbaijan’ genotypes, respectively. Likewise, this PGR combination provided the shortest time until bud break (approximately 6.5 days) and root emergence (approximately 10 days) in both genotypes. The highest number of shoots (4.78 per explant) and roots (3.96) was achieved in this medium in the ‘Kashan’ rose. Stem and root lengths, as well as stem and root fresh and dry weights, were also analyzed. In most measured traits, the lowest values were found in the PGRs-free control medium. Rooted plantlets were transferred to pots filled with perlite and peat moss in a 2:1 proportion and were acclimatized to ambient greenhouse conditions with a mean 90.12% survival rate. This research contributes significantly to our understanding of Damask rose propagation and has practical implications for the cosmetic and ornamental plant industries. By offering insights into the manipulation of regeneration processes, our study opens up new possibilities for the effective production of high-quality plant material. Full article

Ficus palmata is an important fig species that produces edible and nutritious fruit and possesses several therapeutic uses. This study reports an effective method for the micropropagation of F. palmata using nodal explants. In vitro shoots were cultured for 7 weeks onto MS medium fortified with different concentrations of cytokinins, light intensities, sucrose concentrations, and light/dark incubation treatments. Optimal axillary shoot proliferation (10.9 shoots per explant) was obtained on a medium containing 30 g/L sucrose and supplemented with 2 mg/L 6-benzylaminopurine (BAP) under 35 μmol/m²/s light intensity. Dark incubation limited the foliage growth but favored shoot elongation and rooting compared with light incubation. Elongated shoots, under dark conditions, were rooted (100%; 6.67 roots per explant) onto MS medium containing 1 mg/L indole-3-acetic acid (IAA) and 1.5 g/L activated charcoal. The micropropagated plantlets were acclimatized with a 95% survival rate. In this study, the genetic fidelity of micropropagated F. palmata clones along with their mother plant was tested using randomly amplified polymorphic DNA (RAPD), inter-simple sequence repeats (ISSR), and start codon targeted (SCoT) molecular markers. The genetic similarity between the micropropagated plantlets and the mother plant of F. palmata was nearly 95.9%, assuring high uniformity and true-to-type regenerated plants. Using micropropagated F. palmata plantlets as a rootstock proved appropriate for the grafting F. carica ‘Brown Turkey’. These findings contribute to the commercial propagation and production of the fig crop. Full article

Plants’ genetic improvement continues to be crucial for modern agriculture, while biotechnology can offer efficient tools that enhance the selection and recommendation processes of elite clones. This work established a suitable methodology for the regeneration of blueberry (Vaccinium corymbsum) plants in cultures with colchicine. This could be considered a basis for producing populations for the selection of clones following a genetic improvement program assisted by biotechnology. The factors studied were: (a) explant type (leaf discs; nodal segments); (b) colchicine concentration (0, 0.5, 1, and 2 mg/L); and (c) time of exposure to colchicine (1, 2, 3, 5, and 30 days). The basal medium McCown’s Woody Plant (WP) supplemented with 2 mg/L 2iP and 1 mg/L BAP was used with the commercial genotype Duke as a model. A total of 1957 blueberry clones were produced in a medium with 1 mg/L colchicine, distributed at different exposure times. Flow cytometry analyses revealed the following patterns: single patterns for random samples of control plants (Duke donor) and some clones regenerated on colchicine; double patterns for chlorotic plants regenerated on colchicine. Triple and quadruple patterns were observed in callus tissues that did not regenerate plants on colchicine. Populations of plants regenerated in colchicine (6787) and control plants regenerated in in vitro culture without colchicine were adapted under greenhouse conditions. The variables evaluated at this stage were adaptability, height, diameter, number of leaves, incidence of diseases, flowering capacity, and agrobotanical traits. Selected clones demonstrating phenotypic variability (157 clones) were transplanted to field conditions. From the clonal field trial conducted under minimum tillage conditions, 38 clones were selected for improved traits related to the agricultural yield and nutritional quality of the fruits. Of these, six clones showed the highest agronomic performance and adaptability to adverse environmental conditions compared to the Duke donor genotype. It is recommended that these clones continue genotype × environment interaction trials at different locations. Full article

Industrial hemp Cannabis sativa L. is an economically important crop mostly grown for its fiber, oil, and seeds. Due to its increasing applications in the pharmaceutical industry and a lack of knowledge of gene functions in cannabinoid biosynthesis pathways, developing an efficient transformation platform for the genetic engineering of industrial hemp has become necessary to enable functional genomic and industrial application studies. A critical step in the development of Agrobacterium tumefaciens-mediated transformation in the hemp genus is the establishment of optimal conditions for T-DNA gene delivery into different explants from which whole plantlets can be regenerated. As a first step in the development of a successful Agrobacterium tumefaciens-mediated transformation method for hemp gene editing, the factors influencing the successful T-DNA integration and expression (as measured by transient β-glucuronidase (GUS) and Green Florescent Protein (GFP) expression) were investigated. In this study, the parameters for an agroinfiltration system in hemp, which applies to the stable transformation method, were optimized. In the present study, we tested different explants, such as 1- to 3-week-old leaves, cotyledons, hypocotyls, root segments, nodal parts, and 2- to 3-week-old leaf-derived calli. We observed that the 3-week-old leaves were the best explant for transient gene expression. Fully expanded 2- to 3-week-old leaf explants, in combination with 30 min of immersion time, 60 µM silver nitrate, 0.5 µM calcium chloride, 150 µM natural phenolic compound acetosyringone, and a bacterial density of OD_600nm = 0.4 resulted in the highest GUS and GFP expression. The improved method of genetic transformation established in the present study will be useful for the introduction of foreign genes of interest, using the latest technologies such as genome editing, and studying gene functions that regulate secondary metabolites in hemp. Full article