Search Results (58)

Genetic transformation is an essential tool for investigating gene function and editing genomes. Kiwifruit, recognized as a significant global fresh fruit crop, holds considerable economic and nutritional importance. However, current genetic transformation techniques for kiwifruit are impeded by low efficiency, lengthy culture durations (a minimum of six months), and substantial labor requirements. In this research, we established an efficient system for shoot regeneration and the stable genetic transformation of the ‘Hayward’ cultivar, utilizing leaf explants in conjunction with two strains of Agrobacterium that harbor the expression vector pBI121-35S::GFP, which contains the green fluorescent protein (GFP) gene as a visible marker within the T-DNA region. Our results show that 93.3% of leaf explants responded positively to the regeneration medium, producing multiple independent adventitious shoots around the explants within a six-week period. Furthermore, over 71% of kanamycin-resistant plantlets exhibited robust GFP expression, and the entire transformation process was completed within four months of culture. Southern blot analysis confirmed the stable integration of GFP into the genome, while RT-PCR and fluorescence microscopy validated the sustained expression of GFP in mature plants. This efficient protocol for regeneration and transformation provides a solid foundation for micropropagation and the enhancement of desirable traits in kiwifruit through overexpression and gene silencing techniques. Full article

Melastoma dodecandrum is primarily propagated through stem cuttings, which limits genetic variation and constrains breeding efforts. To overcome this limitation and facilitate molecular breeding, the establishment of a reliable and efficient regeneration system is essential. This study investigated the effects of plant growth regulators (PGRs) and culture media on the in vitro regeneration system of M. dodecandrum. The highest rate of callus induction (96.67%) was achieved when sterile leaf explants were cultured on Murashige and Skoog (MS) basal medium supplemented with 2.00 mg·L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.50 mg·L⁻¹ 6-benzylaminopurine (6-BA). For callus differentiation, the optimal formulation of MS + 2.0 mg·L⁻¹ 6-BA + 0.5 mg·L⁻¹ naphthylacetic acid (NAA) resulted in a differentiation frequency of 83.33%. The optimal PGR combinations for shoot proliferation were 1.5 mg·L⁻¹ 6-BA + 0.1 mg·L⁻¹ NAA and 0.5 mg·L⁻¹ 6-BA + 0.2 mg·L⁻¹ NAA. The optimal rooting media were MS medium supplemented with 0.1, 0.2, or 0.5 mg·L⁻¹ indole-3-butyric acid (IBA) or 1/2MS medium supplemented with 0.1 mg·L⁻¹ IBA. Additionally, this study investigated the dynamic changes in endogenous hormones during the regeneration process. The levels and ratios of hormones, including gibberellin (GA₃), abscisic acid (ABA), indole-3-acetic acid (IAA), and zeatin (ZT), collectively regulated the regeneration process. Elevated levels of ABA and GA₃ may promote callus initiation as well as the growth and development of adventitious roots during the early induction stage. Reduced levels of ABA and IAA favored callus differentiation into shoots, whereas elevated GA₃ levels facilitated proliferation of adventitious shoots. Throughout the regeneration process, fluctuations in ZT levels remained relatively stable. This study successfully established an in vitro regeneration system for M. dodecandrum using leaf explants, providing theoretical guidance and technical support for further molecular breeding efforts, genetic transformation, and industrial development. Full article

Light influence on shoot regeneration in Prunus salicina is a complex interaction that has been studied for the first time. Japanese plum plants were regenerated from calli and seeds of the scion cultivar ‘Victoria’. The effect of four different light spectra (white, blue, red, and mixed), along with three 6-benzyladenine (BA) concentrations (1, 1.5, and 2 mg L⁻¹), was studied in these two sources of explants. Organogenic calli were derived from the base of stem explants of the scion cultivar ‘Victoria’, whereas cotyledons and embryogenic axis slices were used as seed explants. Calli cultured with 2 mg L⁻¹ of BA and mixed light or 2.5 mg L⁻¹ of BA and control light showed the highest regeneration rates, with no significant differences compared to other treatments. Seed explants exposed to 2.5 mg L⁻¹ of BA and red light exhibited significantly higher organogenesis. In comparison, those in 1.5 mg L⁻¹ of BA with blue light or 2.5 mg L⁻¹ of BA with mixed/control light showed no regeneration. BA concentration did not have a significant effect in the induction of somatic shoots from any explant source. In contrast, a strong interaction between light and BA was noticed. This work presents a protocol that can be applied in transformation and editing research as light spectrum studies continue to advance. Full article

Bombax ceiba is an important medicinal and ornamental tree widely distributed in tropical and subtropical areas. However, its seeds lose viability rapidly after harvest, which has created hurdles in large-scale propagation. Here, we describe the development of a rapid and efficient de novo organogenesis system for Bombax ceiba, incorporating both indirect and direct regeneration pathways. The optimal basal medium used throughout the protocol was ½ MS supplemented with 30 g/L glucose, with all cultures maintained at 26–28 °C. For the indirect pathway, callus was induced from both ends of each hypocotyl on basal medium supplemented with 0.2 mg·L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5 mg·L⁻¹ 6-Benzylaminopurine (6-BA) under dark conditions. The induced calluses were subsequently differentiated into adventitious shoots on basal media containing 0.5 mg·L⁻¹ Indole-3-butyric acid (IBA), 0.15 mg·L⁻¹ Kinetin (KIN), and 1 mg·L⁻¹ 6-BA under a 16 h photoperiod, resulting in a callus induction rate of 140% and a differentiation rate of 51%. For the direct regeneration pathway, shoot buds cultured on medium with 0.5 mg·L⁻¹ IBA and 1 mg·L⁻¹ 6-BA achieved a 100% sprouting rate with a regeneration coefficient of approximately 3.2. The regenerated adventitious shoots rooted successfully on medium supplemented with 0.5 mg·L⁻¹ Naphthylacetic acid (NAA) and were acclimatized under greenhouse conditions to produce viable plantlets. This regeneration system efficiently utilizes sterile seedling explants, is not limited by seasonal or environmental factors, and significantly improves the propagation efficiency of Bombax ceiba. These optimized micropropagation methods also provide a robust platform for future genetic transformation studies using hypocotyls and shoot buds as explants. 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

Philodendron ‘White Knight’ is a popular climbing evergreen plant typically propagated through stem cuttings. However, this method is slow and inefficient, making it challenging to meet the rising market demand. In vitro propagation could enhance the multiplication of this cultivar. However, research on its in vitro propagation is limited. Therefore, the objective of the present study was to establish an efficient micropropagation technique to mass-produce Philodendron ‘White Knight’ to meet the market demand. We investigate the impact of silver nanoparticles (Ag NPs) on the surface sterilization of Philodendron ‘White Knight’ petioles, the role of plant growth regulators in adventitious shoot regeneration and shoot multiplication, and the effect of auxins on the rooting ability of Philodendron ‘White Knight’ microshoots. There are few stages in plant micropropagation. The establishment of aseptic culture is the first and most important stage. For Philodendron ‘White Knight’, aseptic petiole explants (100%) were obtained after treatment with 40 mg L⁻¹ Ag NPs for 60 min. This was followed by adventitious shoot induction, and the highest rate of adventitious shoot induction (52.6%) and the maximum shoot number (13.9 shoots per petiole) were achieved on Murashige and Skoog shoot multiplication B (MS-B) medium with 20 µM of 2-isopentenyl adenine (2-IP) and 5.0 µM of naphthalene acetic acid (NAA). The shoot multiplication stage was achieved with the highest number of shoots (34 shoots per shoot tip) with a length of 5.1 cm, which was obtained on MS-B medium with 5.0 µM 2-IP and 2.5 µM NAA. All the microshoots produced roots during the root induction stage with the maximum root number (8.2 roots per shoot), and the greatest plantlet height (9.1 cm) was achieved on half-strength Murashige and Skoog medium containing indole-3-butyric acid (10.0 μM). The rooted plantlets of Philodendron ‘White Knight’ were transplanted into a substrate composed of 10% peat moss, 50% orchid stone, and 40% coconut husk chips and acclimatized in a greenhouse environment, achieving a survival rate of 100%. This micropropagation protocol can be used for the commercial production of Philodendron ‘White Knight’. Full article

Populus species are important resources for ecological conservation and certain industry productions, and are also considered model tree species for scientific research. For tree species, in vitro plant regeneration is an important method of propagation due to the advantage of high multiplication rate. Although many molecular determinants for poplar regeneration have been investigated, the complete regulatory hierarchy network remains unclear. In this study, we tracked the temporal changes of endogenous hormone contents, physiological characteristics and transcriptional profiles during callus induction and adventitious organogenesis in a stem of Populus alba L. to explore the regulatory dynamics of in vitro regeneration in poplars. The results imply that auxin may promote the formation of callus in P. alba by activating the expression of WOX11/12. By up-regulating the expression of CUC1/2, the development of callus begins to initiate apical meristem (SAM) at day 12. The cytokinin-mediated pathway regulates the adventitious shoot formation by ESR1 and WUS. The precursors of active gibberellin GA1, GA53 and GA19 were accumulated in the early stage of callus induction, and then they continued to decrease. JA may function on adventitious shoot regeneration due to its accumulation after 12 days of induction. The dominant hormonal components and regulatory factors during regeneration were identified. Based on the results, a regeneration pathway regulated by auxin and cytokinin for poplars is proposed. The key regulators identified in this study will accelerate the exploration and understanding of the asexual reproduction mechanism of poplar trees. Full article

Quercus robur L., also referred to as “summer oak” or “English oak”, is an esthetically pleasing species, making it an excellent choice for street trees and gardens. Raising Quercus presents several challenges, including its long growth period, delayed germination, and inconsistent emergence. The shoot proliferation and adventitious root formation of Q. robur are crucial for establishing a tissue culture regeneration system and are vital for the successful transplantation of seedlings. To address this, experiments were conducted to assess shoot proliferation and adventitious root formation in Q. robur using various media. The shoot proliferation time, shoot proliferation coefficient, number of rooting strips, and length indicators of roots were recorded. The results indicated that a combination of 0.3 mg/L 6-Benzylaminopurine (6-BA) and 100 mg/L cefotaxime (Cef) was optimal for shoot propagation, while a solution of 0.1 mg/L 1-Naphthaleneacetic acid (NAA) and 1/2 Murashige and Skoog Medium (1/2MS) medium was most effective for root induction. This study has identified the optimal conditions for adventitious root formation and shoot proliferation in Q. robur, providing a basis for further research into propagation, germplasm conservation and genetic transformation techniques. Full article

Blueberries are a relatively recently domesticated species, primarily bred through hybridization. Mutation breeding, which uses chemical or physical treatment to increase plant mutation, has not yet been applied to blueberries. This study introduces a mutation breeding strategy for the highbush blueberry cultivar Vaccinium corymbosum. We established a high-efficiency regeneration protocol, which was applied to leaves and stems exposed to gamma irradiation using ⁶⁰Co-γ rays at doses of 10, 20, 40, 80, and 120 gray (Gy), to increase the efficiency of mutated cells to develop into adventitious shoots. We determined that the median lethal dose (LD₅₀) was approximately 56 Gy for leaf explants and 80 Gy for stem explants. Phenotypic variations, including changes in leaf color and growth characteristics, which may be due to altered plant response to environmental factors, were successfully observed in the first-generation (M1) plants. The height of M1 plants quantitatively decreased with increasing irradiation doses. To evaluate the mutants induced by each irradiation dose, whole-genome resequencing was conducted on individuals from each dose group, revealing significant genomic alterations at the 80 Gy dose. This approach provides a valuable reference for future blueberry breeding programs aimed at enhancing genetic diversity and improving cultivar performance. Full article

Haworthia truncata, a species native to South Africa, is characterized by its limited growth and scarcity, contributing to high production costs. Countries like China and Turkey are known for exporting Haworthia globally. Tissue culture offers an efficient method for mass-producing unique and beautiful species such as H. truncata. This study tested Murashige and Skoog (MS) basal media supplemented with various concentrations of IBA (0.05–1.5 mg/L), NAA (0.05–0.25 mg/L), and BA (0.25–1.5 mg/L) to promote shoot proliferation. MS medium without plant growth regulators (PGRs) was also tested as a control. Different explant types (leaf, root, and inflorescence) were analyzed for their potential in direct and indirect regeneration. Inflorescence explants showed the highest callus induction with 1.5 mg/L IBA, while optimal shoot proliferation occurred at 1 mg/L IBA. Callus induction was optimal for leaf explants with 0.05 mg/L NAA and 0.25 mg/L BA, and shoot proliferation was highest at 0.05 mg/L NAA and 1 mg/L BA. Root explants achieved maximum callus induction with 0.25 mg/L BA and 0.25 mg/L NAA, with the best shoot proliferation using 0.05 mg/L NAA and 1 mg/L BA. The highest rooting percentage of regenerated shoots was obtained on ½ MS medium with 1.5 mg/L IBA. Full article

Kalanchoe beharensis, a vulnerable species according to the International Union for Conservation of Nature, is highly prized for its ornamental value and medicinal properties. Therefore, an efficient methodology to propagate this ecologically significant species would be of particular interest. The propagation of K. beharensis has traditionally been achieved by seed or cuttings, but these methods are limited in efficiency. Micropropagation provides a more efficient and controlled alternative by enabling the in vitro production of numerous plants in a small space and in a short period of time. Despite its advantages, no micropropagation protocol for K. beharensis has been reported in the literature. In this study, we report an efficient in vitro regeneration protocol for K. beharensis. In order to implement this, we evaluated the morphogenetic response of leaf and root explants in media supplemented with auxins, cytokinins, or a combination of both growth regulators. Surprisingly, the best results were observed in indole-3-acetic acid-supplemented media. Adventitious shoots were rooted in either hormone-free or auxin-supplemented media, with indole-3-acetic acid yielding the best results. Rooted plants were acclimatized in the greenhouse, achieving over 80% survival during acclimatization. This protocol improves multiplication rate, space utilization, and uniformity, providing a viable alternative to conventional propagation methods. Full article

Paeonia ostii, a plant of substantial economic significance, continues to face constraints in achieving large-scale propagation. In vitro propagation offers a promising avenue for the production of disease-free plants and the genetic transformation of peonies to instill novel traits. However, significant challenges persist in tissue culture, particularly with regards to the reproduction coefficient of shoots and the rooting process. This study reports an efficacious protocol for P. ostii micropropagation, focusing on in vitro root development facilitated through the application of phloroglucinol (PG). Furthermore, the study unveils the molecular signature of P. ostii during in vitro root development. The results indicate that the modified Y3 medium (Y3M), supplemented with 1 mg/L 6-benzyladenine (BA) and 0.1 mg/L α-naphthaleneacetic acid (NAA), is optimal for adventitious bud induction, achieving a 96.67% induction rate and an average of 16.03 adventitious shoots per sample. The highest elongation percentage (92.15%) and the longest average shoot length (3.87 cm) were obtained with Y3M containing 0.3 mg/L BA and 0.03 mg/L NAA. Additionally, the optimal medium for inducing root formation in P. ostii was identified as WPM supplemented with 3 mg/L indole-3-butyric acid (IBA) and 100 mg/L phloroglucinol (PG). Lignin content detection, microscope inspection, and molecular signature results demonstrated that PG enhanced lignin biosynthesis, thereby promoting in vitro rooting of P. ostii. Full article

Mango (Mangifera indica L.) is one of the most significant tropical and subtropical fruit species, with high ecological and economic value. However, research on the in vitro culture of mangoes is relatively weak, so establishing an efficient and stable mango plant regeneration system is of great significance. In this study, a preliminary mango regeneration system was established with Mangifera indica L. cv. Keitt from young branches as the starting explants. The results showed that the optimal plant growth regulator (PGR) formula for direct adventitious shoot induction on the branches was 1 mg/L 6-benzylaminopurine (6-BA) + 0.1 mg/L a-naphthaleneacetic acid (NAA), with an adventitious shoot induction rate of 73.63% and an average of 6.76 adventitious shoots. The optimal basal medium for adventitious shoot induction was wood plant medium (WPM), with an adventitious shoot induction rate of 63.87% and an average of 5.21 adventitious shoots. The optimal culture medium for adventitious shoot elongation was WPM + 1 mg/L 6-BA + 0.5 mg/L NAA, with an adventitious shoot elongation rate of 89.33% and an average length of 5.17 cm. The optimal formula for the induction of mango rooting was Douglas fir cotyledon revised medium (DCR) + 3 mg/L indole-3-butyric acid (IBA), with a maximum rooting rate of 66.13% and an average rooting quantity of 6.43. The genetic fidelity of the in vitro-regenerated plants was evaluated using inter-simple sequence repeat (ISSR) molecular markers. There was no difference between the in vitro-regenerated plants and the parent plant. This study provides an efficient and stable propagation system for Mangifera indica L., laying the foundation for its rapid propagation and genetic improvement. Full article

Allium microdictyon Prokh. is a rare, endemic species possessing good taste qualities and listed in the Red Book of Kazakhstan; therefore, it is subject to anthropogenic impact (food gathering, grazing, logging, fires, etc.), which leads to a substantial reduction of its area. The aim of the study was to develop a protocol for microclonal propagation of A. microdictyon. Mature seeds of A. microdictyon collected from natural habitats in the Kazakhstani Altai were used as explants. Optimization of seed sterilization methods, selection of growth regulators for inducing adventitious shoot formation and microclonal propagation, and optimization of conditions for adaptation of regenerants to ex vitro conditions were carried out. Surface sterilization of seeds with 70% EtOH and 0.01% HgCl₂ is optimal for obtaining sterile and viable A. microdictyon seedlings. Sterile seedlings obtained in vitro on ½ Murashige and Skoog medium supplemented with 10 mg L⁻¹ gibberellic acid and 0.1 mg L⁻¹ indole-3-acetic acid (IAA) were used as a source for obtaining micropropagation cultures. Induction of adventitious organogenesis of A. microdictyon was effective on media containing 0.5 mg L⁻¹ 6-benzylaminopurine (BAP) and 1.5–2 mg L⁻¹ zeatin. On these variants, leaf conglomerates consisting of abundantly overgrown thin leaves were formed. The effect of 0.2 mg L⁻¹ indole-3-butyric acid (IBA) on further development of organogenesis and formation of microbulbs in A. microdictyon was shown in comparison with IAA, NAA, and PAC. Regenerated A. microdictyon plants were adapted to ex vitro conditions and resumed growth after 16–20 weeks of relative dormancy. The developed micropropagation protocol can be used to preserve germplasm and propagate for subsequent restoration of A. microdictyon populations in natural habitats. Full article

Chrysanthemum morifolium ‘11-C-2’ is a variety of chrysanthemums with high ornamental and tea value, experiencing significant market demand. However, as cultivation areas expand, issues such as viral infection, germplasm degradation, low proliferation coefficient, and slow proliferation rate arise, necessitating the establishment of an efficient in vitro regeneration system. This study, based on the principles of orthogonal experimental design, explored the regeneration system of Chrysanthemum cultivar ‘11-C-2’ using sterile seedlings. The research focused on three key stages: adventitious bud differentiation, rooting culture, and acclimatization–transplantation, employing shoot-bearing stem segments and leaves as explants. The findings indicate that the optimal explant for the Chrysanthemum ‘11-C-2’ sterile seedlings is the shoot-bearing stem segment. The best medium for adventitious bud differentiation was determined to be MS supplemented with 1.5 mg/L 6-BA and 0.5 mg/L NAA. Bud differentiation began on day 17 with a 100% differentiation rate, completing around day 48. The maximum differentiation coefficient reached 87, with an average of 26.67. The adventitious buds were then cultured for rooting in the optimal medium of 1/2 MS supplemented with 0.1 mg/L NAA. Rooting was initiated on day 4 and was completed by day 14, achieving a rooting rate of 97.62%. After a 5-day acclimatization under natural light, the rooted seedlings were transplanted into a growth substrate with a peat-to-vermiculite ratio of 1:2. The plants exhibited optimal growth, with a transplantation survival rate of 100%. The findings provide data support for the efficient large-scale propagation of ‘11-C-2’ and lay the foundation for germplasm preservation and genetic transformation research of tea chrysanthemums. Full article