Search Results (39)

Propagation of hinoki cypress (Japanese cypress, Chamaecyparis obtusa, Cupressaceae) through adventitious bud multiplication was performed using leaf-segment explants from cutting plants of selected adult trees. Explants were successfully surface-sterilized (>90% asepsis) by agitating them in 2.5% (w/v available chlorine) sodium hypochlorite solution for 15 min and then rinsed with sterile distilled water. Explants approximately 2 cm long were cultured on plates containing medium supplemented with 6-benzylaminopurine (BAP) and 2,4-dichlorophenoxyacetic acid (2,4-D), 20 g/L sucrose, and 7 g/L agar. The cultures were kept at 25 ± 1 °C under a 16-h photoperiod with a photon flux density of approximately 65 µmol m⁻² s⁻¹. The optimal adventitious bud multiplication (31.5 buds per explant) was obtained on a medium supplemented with 10 µM BAP in combination with 1 µM 2,4-D. Proliferated adventitious buds were elongated better on medium supplemented with 1 µM trans-zeatin. The best rooting result (86%) was achieved on a rooting medium supplemented with 1 µM 3-indolebutyric acid in combination with 0.1 µM 1-naphthaleneacetic acid. However, rooting response varied according to genotypes. Clones related to the cultivar ‘Nangouhi’ (Na18, Na14 x Isa, Na14-14, Isa x Na14, and NaS) were easier to root than those derived from the cultivar ‘ShizuokaKenZairai’ (SKZ5 and SKZ8). Regenerated plantlets did not show morphological abnormalities and showed a high survival rate after acclimatization (>90%). Full article

Two bulbous plant species, Squilla maura and Oncostema peruviana (Asparagaceae), show particularly interesting ornamental and medicinal characteristics. Micropropagation could be a promising alternative method to accelerate their naturally slow spreading pattern. This study focused on the effects of different growth regulators and culture media on callus induction and shoot regeneration, from which an effective protocol was established. Leaf explants were cultured on Murashige and Skoog (MS), MS/2, or B5 medium, containing one auxin—1-Naphthaleneacetic acid (NAA), 2,4-Dichlorophenoxyacetic acid (2.4-D), or 3-indolebutyric acid (IBA)—combined with the cytokinin 6-Benzylaminopurine (BAP), at various concentrations. All treatments resulted in callogenesis rates between 65–100% for both species. These treatments led to direct bulbil regeneration. Among the treatments, the B5 medium with 1 mgL⁻¹ BAP and 1 mgL⁻¹ 2,4-D gave the highest regeneration rate (89.2%) for O. peruviana, while the ½MS medium with 0.5 mgL⁻¹ BAP and 0.5 mgL⁻¹ NAA showed the highest regeneration rate (85.5%) for S. maura. The highest mean number of bulils per explant was 7.44 for O. peruviana on the MS medium (0.5 mgL⁻¹ BAP and 2 mgL⁻¹ IBA), and 9.5 for S. maura on the MS medium (1 mgL⁻¹ BAP and 0.5 mgL⁻¹ NAA). The regenerated bulbils were transferred for multiplication to the MS medium with a hormone combination (2 mgL⁻¹ BAP and 0.2 mgL⁻¹ NAA) which increased the multiplication rate compared to the callus induction me dium, with a highest recorded multiplication rate of 177 (O. peruviana) and 104.33 (S. maura). The propagation stage achieved the highest number of bulbils/explant after a second subculture for the two species. An efficient micropropagation protocol was established for S. maura, which answers our main objective, and it would contribute to their conservation and sustainable use in ornamental and medicinal applications. Full article

The Duboisia species, a group of plants native to Australia, have been historically valued for their pharmacological properties and have played a significant role in traditional medicine and pharmaceutical research. Persistent efforts are underway to enhance the efficacy of the active ingredient scopolamine, employing both conventional breeding methods and advanced biotechnology tools. The primary objective of this research was to establish a highly efficient method for isolating mesophyll protoplasts and facilitating their regeneration, thereby laying a robust foundation for the application of various advanced plant biotechnology tools in the pursuit of genetic enhancement. The mesophyll protoplast isolation process was developed for hybrid D. myoporoides × D. hopwoodii with careful optimisation of the following parameters: leaf strip size; incubation conditions; physical treatment; and enzyme concentration. The optimal parameters were combined in each individual step; the best enzyme concentration was determined to be 2% (w/v) cellulysin and 0.5% (w/v) macerase. Protoplast yield was found to be greatly affected by the enzyme concentrations. The isolated protoplasts were cultured at a density of 0.5 × 10⁵ to best sustain the highest cell division (33.2%) and a microcalli induction frequency of 17.9%. After 40 days of culture in a modified KM8P medium at 25 °C in darkness, visible microcalli were transferred to a solidified Murashige and Skoog (MS) medium with 1 mg L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) for callus induction under a 16 h photoperiod. After 30 days of culture, compact organogenic calli were transferred into a solid MS medium with 6-benzylaminopurine (BA) alone or thidiazuron (TDZ) alone or in combination with BA or naphthalene acetic acid (NAA) for shoot regeneration. The maximum shoot regeneration frequency (63.3%) was observed in the medium with 1.5 mg L⁻¹ TDZ alone. For the first time, a reliable protoplast isolation and regeneration system from mesophyll cells was established for Duboisia with high protoplast viability, successful microcalli formation, and intact plant regeneration. This innovation will significantly contribute towards the genetic enhancement of the Duboisia species. Full article

Opuntia is recognized economically as a significant crop for its nutritional, functional, and therapeutic properties and its potential in the pharmaceutical, cosmetic, and bioenergy industries. Opuntia is conventionally propagated by seed or vegetative propagation via rooted scions or grafting. However, multiplication procedures are insufficient for extensive spreading. One of the alternative techniques is in vitro. For this reason, the objective was to evaluate the growth of different components combined with light conditions to induce callus, embryogenesis, and the possible synthesis of a component in the genus Opuntia. The existence of genes involved in pigment synthesis in genotypes of different fruit colors was complementarily analyzed. In this study, we used different combinations of fructose (2 g/L) with prickly pear juice (2 mL/L), mannitol (4 g/L), silver nitrate (AgNO₃: 1 mg/L), 2,4-dichlorophenoxyacetic acid (2,4-D: 2 mg/L), and indole-3-acetic acid (IAA: 1 g/L), with white, blue, and red LEDs (light-emitting diodes) and laser beams. These explants yielded sufficient contents of simple phenols, gallic acid equivalents (GAE: 2283.30 ± 6.29 mg/100 g), and coumaric (2155.0 ± 35.0 mg/100 g) and ferulic (2176 ± 27.9 mg/100 g) acids for the genotype Tapón aguanoso, and chlorogenic acid (CGA: 380.22 ± 22.05 mg/100 g) for the Copena V1 genotype. Upon analyzing the genotypes of different fruit pigmentations, we also observed the following: enzyme 4,5 estradiol dioxygenases can be present in all genotypes (regardless of pigmentation); enzyme 5,3 glucosyltransferases (GTs) could be induced in pigment synthesis in the genotypes of orange to purple fruits. Sequencing primer-amplified fragments for GT showed high similarity to uridine diphospho (UDP)-glucose from other species. This allows us to infer that it is possible to obtain products of high therapeutic value in the near future under controlled conditions. Full article

Nagpur mandarin is a popular table fruit across India and is exported to various countries. Only 1% of the total production is processed into various products. The development of seedless cultivars will boost agricultural incomes by enhancing the potential for export and processing. At present, only a couple of commercially seedless varieties are available, but these have yet to become popular. A research study was undertaken to quickly develop a high-quality seedless variety of Nagpur mandarin by combining the available technologies, viz., endosperm rescue, somatic embryogenesis, and mini-grafting, at CCRI, Nagpur. Complete plantlets of C. reticulata Blanco cv. Nagpur mandarin was successfully regenerated from hybrid endosperm tissue via somatic embryogenesis after attempting various permutations and combinations of media at various stages of regeneration, right from the primary callus to complete plantlet production. Maximum response (93.33%) and survival (91.67%) for primary callus induction were obtained in Murashige and Tucker (MT) + Malt Extract (ME) + Casein Hydrolysate (CH) (500 mg/L) +2,4-dichlorophenoxyacetic acid (2,4-D) (2 mg/L) medium. Maximum stimulation for embryogenesis and morphogenesis occurred in 2MT + CH (500 mg/L) + adenine sulfate (ad.s) (2 mg/L) + Benzyl Adenine (BA) (0.25 mg/L). The highest response (95.84%) for shoot differentiation occurred in MT + adenine sulphate (2 mg/L) + Gibberellic Acid (GA3) (1 mg/L) + BA (1 mg/L) (94.85%). The surviving plantlets were tested for ploidy status through flow Cytometry, chromosomal counting/cytogenetic technique, leaf morphology, stomatal characteristics, and the appearance of prominent thorns. In initial evaluation trials, the fruits of the triploid field-planted trees were found to be commercially seedless. These results demonstrated the recovery of stable triploids from the hybrid endosperm via somatic embryogenesis, which is the first of its kind in the field of Citrus triploid breeding in India. Full article

Rice blast caused by Magnaporthe oryzae is one of the most serious rice diseases worldwide. The early indica rice thermosensitive genic male sterile (TGMS) line HD9802S has the characteristics of stable fertility, reproducibility, a high outcrossing rate, excellent rice quality, and strong combining ability. However, this line exhibits poor blast resistance and is highly susceptible to leaf and neck blasts. In this study, backcross introduction, molecular marker-assisted selection, gene chipping, anther culture, and resistance identification in the field were used to introduce the broad-spectrum blast-resistance gene R6 into HD9802S to improve its rice blast resistance. Six induction media were prepared by varying the content of each component in the culture medium. Murashige and Skoog’s medium with 3 mg/L 2,4-dichlorophenoxyacetic acid, 2 mg/L 1-naphthaleneacetic acid, and 1 mg/L kinetin and N6 medium with 800 mg/L casein hydrolysate, 600 mg/L proline, and 500 mg/L glutamine could improve the callus induction rate and have a higher green seedling rate and a lower white seedling rate. Compared to HD9802S, two doubled haploid lines containing R6 with stable fertility showed significantly enhanced resistance to rice blast and no significant difference in spikelet number per panicle, 1000-grain weight, or grain shape. Our findings highlight a rapid and effective method for improving rice blast resistance in TGMS lines. Full article

Fritillaria meleagris is a horticulturally and medicinally valuable bulbous plant that requires a period of low temperatures for proper growth and flowering. Since conventional methods of propagation are ineffective and very slow, tissue culture techniques offer an integrated approach to mass production of this valuable geophyte. In this study, we investigated the effects of various auxin–cytokinin combinations on different morphogenetic pathways in bulb scale culture. Bulbs obtained in vitro were cut longitudinally, and bulb scales were cultured for four weeks at 7 °C on MS medium supplemented with 6-benzylaminopurine (BAP) in combination with 2,4-dichlorophenoxyacetic acid (2,4-D) or α-naphthaleneacetic acid (NAA) at different concentrations in order to investigate the influence of plant growth regulators (PGRs) on different morphogenetic responses. Regeneration percentage, number of shoots per explant, shoot length, number of bulbs and number of somatic embryos were monitored weekly. After chilling, bulb scales were transferred to 24 °C, and all parameters were recorded again. Low PGR concentrations were very effective for shoot multiplication, yielding up to 5.5 shoots per explant. 2,4-D (at 2 mg/L) in combination with low BAP (0.25 mg/L) produced the highest number of bulbs (11.00 ± 0.00), while PGR-free medium was extremely effective in somatic embryo formation (13.50 ± 2.90). Detached somatic embryos and bulblets continued to grow and develop on fresh PGR-free medium. We present data demonstrating that low auxin–cytokinin concentrations and PGR-free medium provide an effective method for a combined morphogenetic pathway in F. meleagris that is suitable for large-scale propagation. Full article

A superior heterojunction of HC-ZnBi-LDO was synthesized in two steps, namely hydrothermal carbonization, followed by co-precipitation. The 2% HC-ZnBi-LDO heterojunction photocatalysts could degrade over 90.8% of 30 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) using 1.0 g/L of the catalyst after 135 min of visible light exposure at pH 4. The activity of 2% HC-ZnO-LDO was remarkably stable. Approximately 86.4–90.8% of 30 mg/L 2,4-D was degraded, and more than 79–86.4% of TOC was mineralized by 2% HC-ZnBi-LDO at pH 4 after 135 min of visible light exposure during four consecutive cycles. The rapid separation and migration of charge carriers at the interfaces between HC and ZnBi-LDO were achieved within 2% HC-ZnBi-LDO. Moreover, the electron acceptor characteristic of HC in 2% HC-ZnBi-LDO caused the recombination of charge carriers to decrease significantly, thus generating more reactive radicals, such as hydroxyl radicals (OH^●) and superoxide radicals (O₂^●−). These results demonstrate that the novel 2% HC-ZnBi-LDO is a superior photocatalyst for the remediation of hazardous organic pollutants. Full article

This study aimed to establish an efficient plant regeneration system from leaf-derived embryogenic structure cultures of Daphne genkwa. To induce embryogenic structures, fully expanded leaf explants of D. genkwa were cultured on Murashige and Skoog (MS) medium supplemented with 0, 0.1, 0.5, 1, 2, and 5 mg·L⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D), respectively. After 8 weeks of incubation, the highest frequency of embryogenic structure formation reached 100% when the leaf explants were cultivated on MS medium supplemented with 0.1 to 1 mg·L⁻¹ 2,4-D. At higher concentrations of 2,4-D (over 2 mg·L⁻¹ 2,4-D), the frequency of embryogenic structure formation significantly declined. Similar to 2,4-D, indole butyric acid (IBA) and α-naphthaleneacetic acid (NAA) treatments were also able to form embryogenic structures. However, the frequency of embryogenic structure formation was lower than that of 2,4-D. In particular, the yellow embryonic structure (YES) and white embryonic structure (WES) were simultaneously developed from the leaf explants of D. genkwa on culture medium containing 2,4-D, IBA, and NAA, respectively. Embryogenic calluses (ECs) were formed from the YES after subsequent rounds of subculture on MS medium supplemented with 1 mg·L⁻¹ 2,4-D. To regenerate whole plants, the embryogenic callus (EC) and the two embryogenic structures (YES and WES) were transferred onto MS medium supplemented with 0.1 mg·L⁻¹ 6-benzyl aminopurine (BA). The YES had the highest plant regeneration potential via somatic embryo and shoot development compared to the EC and WES. To our knowledge, this is the first successful report of a plant regeneration system via the somatic embryogenesis of D. genkwa. Thus, the embryogenic structures and plant regeneration system of D. genkwa could be applied to mass proliferation and genetic modification for pharmaceutical metabolite production in D. genkwa. Full article

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%. Full article

The conservation of endangered, rare, and endemic plant species is based on in situ and ex situ conservation strategies. When in situ conservation alone is not sufficient to guarantee the survival of the species, ex situ techniques are adopted in support. This study aimed to develop an efficient micropropagation protocol for Adenostyles by evaluating the effect of different plant growth regulators on leaf explants. Adenostyles alpina subsp. macrocephala (Asterace) is a perennial herbaceous plant endemic to Calabria (Southern Italy). The genus Adenostyles includes three species confined to the mountains of the Mediterranean and southern Europe. For callus induction, media supplemented with different concentrations of Benzylaminopurine (BAP) (0.5, 1, 2, and 3 mg L⁻¹), Naphthaleneacetic Acid (NAA) (1 mg L⁻¹), and 2,4-Dichlorophenoxyacetic Acid (2,4-D) (1 mg L⁻¹) were tested. Shoot regeneration and proliferation were obtained in media supplemented with BAP (1, 2, and 3 mg L⁻¹) and NAA (1 mg L⁻¹). Root induction was obtained in media supplemented with IBA (0.25, 0.50, and 1 mg L⁻¹) and NAA (0.25, 0.50, and 1 mg L⁻¹). Statistically significant differences in callus induction and shoot regeneration were observed between the various media tested. The medium containing Murashige and Skoog (MS) supplemented with 3 mg L⁻¹ of BAP and 1 mg L⁻¹ of NAA showed the highest percentage of callus induction and increased shoot regeneration. The regenerated shoots showed more effective root induction in the hormone-free MS medium and in the presence of Indole-3-Butyric Acid (IBA) at concentrations of 0.25, 0.50, and 1 mg L⁻¹. These results can be used as a basis for the preparation of a micropropagation protocol for different taxa of Adenostyles, as well as other species of Asteraceae specialized to the Mediterranean mountain habitat. Full article

Sageretia thea is used in the preparation of herbal medicine in China and Korea; this plant is rich in various bioactive compounds, including phenolics and flavonoids. The objective of the current study was to enhance the production of phenolic compounds in plant cell suspension cultures of Sageretia thea. Optimum callus was induced from cotyledon explants on MS medium containing 2,4-dichlorophenoxyacetic acid (2,4-D; 0.5 mg L⁻¹), naphthalene acetic acid (NAA, 0.5 mg L⁻¹), kinetin (KN; 0.1 mg L⁻¹) and sucrose (30 g L⁻¹). Browning of callus was successfully avoided by using 200 mg L⁻¹ ascorbic acid in the callus cultures. The elicitor effect of methyl jasmonate (MeJA), salicylic acid (SA), and sodium nitroprusside (SNP) was studied in cell suspension cultures, and the addition of 200 µM MeJA was found suitable for elicitation of phenolic accumulation in the cultured cells. Phenolic and flavonoid content and antioxidant activity were determined using 2,2 Diphenyl 1 picrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethybenzothiazoline-6-sulphonic acid (ABTS), ferric reducing antioxidant power (FRAP) assays and results showed that cell cultures possessed highest phenolic and flavonoid content as well as highest DPPH, ABTS, and FRAP activities. Cell suspension cultures were established using 5 L capacity balloon-type bubble bioreactors using 2 L of MS medium 30 g L⁻¹ sucrose and 0.5 mg L⁻¹ 2,4-D, 0.5 mg L⁻¹ NAA, and 0.1 mg L⁻¹ KN. The optimum yield of 230.81 g of fresh biomass and 16.48 g of dry biomass was evident after four weeks of cultures. High-pressure liquid chromatography (HPLC) analysis showed the cell biomass produced in bioreactors possessed higher concentrations of catechin hydrate, chlorogenic acid, naringenin, and other phenolic compounds. Full article

Coffea arabica is one of the two most consumed coffee species in the world. Micropropagation through somatic embryogenesis has allowed the large-scale propagation of different coffee varieties. However, the regeneration of plants using this technique depends on the genotype. This study aimed to develop a protocol for the regeneration of C. arabica L. var. Colombia by somatic embryogenesis for its mass propagation. Foliar explants were cultured on Murashige and Skoog (MS) supplemented with different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (BAP), and phytagel for inducing somatic embryogenesis. In total, 90% of the explants formed embryogenic calli with a culture medium containing 2 mg L⁻¹ of 2,4-D, 0.2 mg L⁻¹ BAP, and 2.3 g L⁻¹ phytagel. The highest number of embryos per gram of callus (118.74) was obtained in a culture medium containing 0.5 mg L⁻¹ 2,4-D, 1.1 mg L⁻¹ BAP, and 5.0 g L⁻¹ phytagel. In total, 51% of the globular embryos reached the cotyledonary stage when they were cultured on the growth medium. This medium contained 0.25 mg L⁻¹ BAP, 0.25 mg L⁻¹ indoleacetic acid (IAA), and 5.0 g L⁻¹ of phytagel. The mixture of vermiculite:perlite (3:1) allowed 21% of embryos to become plants. Full article

The extensive use of 2,4-dichlorophenoxyacetic acid (2,4-D) pesticide leads to the contamination of surfaces and groundwater. In this respect, it is critical to develop an inexpensive and environmentally friendly adsorbent for 2,4-D-laden agricultural leachate. In the current study, termite mound soil (TMS) from Ethiopia was used as an adsorbent in a batch mode aimed at the removal of 2,4-D from an aqueous solution. The TMS was characterized using Brunauer–Emmett–Teller (BET), Fourier-transform infrared (FTIR), scanning electron microscopy (SEM), atomic absorption spectrometry (AAS), and X-ray diffraction (XRD) techniques. The effects of various operating parameters such as pH, contact time, adsorbent dose, and initial concentration were investigated. In addition, the optimization process and interaction effect were studied using response surface methodology (RSM). A high 2,4-D removal percentage (89.6%) was achieved for a 2,4-D initial concentration of 50.25 mg/L at pH 2, an adsorbent dose of 15.25 g/L, and a contact time of 180.5 min. The 2,4-D adsorption isotherms could be adequately described by the Langmuir model (R² = 0.9687), while the kinetics of the 2,4-D adsorption on the TMS best fit the pseudo-second-order model. Overall the study showed that TMS is an effective adsorbent for the removal of 2,4-D from agricultural leachate. Full article

2,4-Dichlorophenoxyacetic acid (2,4-D) is a widely used herbicide worldwide. However, its residues in agricultural products are extremely harmful to human health and to the environment in soil and water. Previous methods for determining 2,4-D in water and soil samples are expensive, cumbersome, and not highly selective. In this study, we developed a novel disposal sensor based on screen-printed electrodes (SPEs) for detecting 2,4-D in wastewater and soil samples. The SPEs were modified with conductive polyaniline (PANI) layer and polyvinyl chloride (PVC) membrane loaded with molecularly printed polymer (MIP). The MIP particles were prepared using 2,4-D as template, methacrylic acid (MAA) as monomer, ethylene glycol dimethacrylate (EGDMA) as cross-linker, and benzoyl peroxide as initiator. The best sensor shows a dynamic concentration range of 10⁻² to 10⁻⁷ M 2,4-D, a detection limit (LOD) of 3.6 × 10⁻⁷ M, Nernst slope (response) of 29.9 mV/decade, and high selectivity over other interfering species previously reported in the literature. The sensors also achieved a short response time of 25 s, high reversibility, and a lifetime of over 2 weeks. The developed sensors were successfully used for determining 2,4-D in real wastewater and soil samples with high accuracy and precision. Full article