Recent Articles

Below are the titles and abstracts from the most recent articles published in In Vitro Cellular and Developmental Biology – Plant. Click on the title to view the full article at Springerlink.


  • Initiation pretreatment with Plant Preservative Mixture™ increases the percentage of aseptic walnut shoots
    on June 30, 2022 at 12:00 am

    Abstract Walnut (Juglans regia L.) is one of the most economically important nut crops and one of the more difficult to micropropagate. Initiation and maintaining contamination-free shoot cultures is difficult for many species especially for woody plants. Cultivars of walnut from field collections exhibited severe microbial and fungal contamination when standard initiation procedure was used. An alternative disinfection procedure was designed and tested to reduce the microbial contamination. One-yr-old shoots of walnut cultivars Kogylnichanu, Milotai 10, Peschansky, and Xin 2 were cut in the field in February 2021. Cuttings of 30 to 40 cm long were surface disinfested and were put into jars with water to force bud expansion. After 2 wk, the swollen dormant buds were excised from the stem, surface disinfected in 0.1% mercuric chloride for 10 min, washed with sterile water, and placed into test tubes with Driver and Kuniyuki medium (DKW) with and without Plant […]

  • A new economical storage technique for strawberry (Fragaria × ananassa Duch.) in vitro
    on June 14, 2022 at 12:00 am

    Abstract Strawberry plants grown in vitro are typically stored and maintained on agar containing Murashige and Skoog (MS) media and sucrose as a carbohydrate source. This method of storing strawberry plants in vitro is expensive and time consuming, requiring sub-culturing onto fresh media every 2 to 3 mo. This study aimed to establish the viability of using a substrate alternative as an economical replacement for MS media, for both long-term storage in vitro and ease of transfer (ex vitro). A protocol was developed for strawberry in vitro using commercially available sterilized peat pellets (Jiffy-7® pellets), to optimize culture conditions and tissue culture practices. Suitability of the peat substrate was measured by the plant’s overall response to culture in vitro and subsequent health and survival ex vitro following deflasking. Included in this study was a comparison of the use of vented vs non-vented tissue culture vessels for their effect on plant […]

  • Efficient and reliable propagation and rooting of foetid juniper (Juniperus foetidissima Willd.), as an endangered plant under in vitro condition
    on June 1, 2022 at 12:00 am

    Abstract Foetid juniper (Juniperus foetidissima Willd.) is one of the most important coniferous species faced with multiple threats and it is difficult to propagate it with sexual reproduction. Therefore, the main goal of this study was finding appropriate salt concentrations and growth regulators for shoot regeneration and rooting of J. foetidissima under in vitro conditions and also shortening time of mass production. The effects of 6 different basal culture media (Olive Medium (OM), Murashige and Skoog (MS), Driver and Kuniyuki (DKW), Woody Plant Medium (WPM), 1/2 MS, and 1/4 MS) on shoot proliferation and two different methods of root induction on explants were investigated. Results revealed that OM culture medium supplemented with 6-benzylaminopurine (BAP) (1 mg L−1) and indole-3-acetic acid (IAA) (0.1 mg L−1) was the best medium for shoot proliferation of J. foetidissima (P < 0.05). The highest percentage of shoot regeneration (100%), […]

  • Rapid transformation and plant regeneration of sorghum (Sorghum bicolor L.) mediated by altruistic Baby boom and Wuschel2
    on June 1, 2022 at 12:00 am

    Abstract The standard stalwart Agrobacterium-mediated transformation protocols for sorghum use differentiating embryogenic callus induced from immature embryos in the genotype BTx430. While reliable, the standard protocols are lengthy, and both time and resource expensive. We have investigated the use of altruistic morphogenic regulator mediated transformation (MRMT) to improve sorghum transformation for use in genome editing. Here, we show a method for rapid transformation and plant regeneration of sorghum (Sorghum bicolor L. cultivar BTx430) mediated by altruistic Baby boom and Wuschel2. Confirming and extending previous recent work using altruistic MRMT, we show that the altruistic MRMT rapid protocol results in a higher transformation frequency expressed as a function of the number of independent events per immature embryos infected and requires less time in a transformation pipeline which translates to an increased number of constructs one full-time equivalent […]

  • Heritable DNA-free genome editing of canola (Brassica napus L.) using PEG-mediated transfection of isolated protoplasts
    on June 1, 2022 at 12:00 am

    Abstract Application of new genome editing technologies for plant improvement depends on efficient delivery of editing components into plant cells. For editing of canola, which is an important oil crop, CRISPR/Cas9 and Agrobacterium-mediated transformation of hypocotyl or stem segments have been the primary methods used. Here, we present the development of an efficient and high-throughput system for protoplast isolation, DNA-free editing, and production of edited canola plants. Protoplast polyethylene glycol (PEG)–mediated transfection was conducted with ribonucleoproteins (RNPs) comprising LbCas12a and a single guide RNA. Edits were found at a frequency of approximately 40% in protoplasts cultured for 2 d, in callus colonies derived from these protoplasts, and in regenerated plants. Out of 30 plants that were regenerated from the protoplasts, 12 were edited. Six of them were with biallelic mutations and another six had monoallelic mutations. Targeted deletions […]

  • Testing two chromosome doubling agents for in vitro tetraploid induction on ginger lilies, Hedychium gardnerianum Shepard ex Ker Gawl. and Hedychium coronarium J. Koenig
    on June 1, 2022 at 12:00 am

    Abstract Polyploidization can be a way to produce new varieties in vegetatively propagated species where options on increasing genetic variability are limited compared with sexual reproduction. While there are hundreds of publications with in vitro methods in somatic doubling, it is cardinal to custom-test for a target species of interest on choosing specific reagents and optimizing conditions. This research was performed to provide a reference process for Zingiberaceae species of which the majority is reproduced with vegetative propagation. Ginger lilies, Hedychium gardnerianum Shepard ex Ker Gawl. and Hedychium coronarium J. Koenig, were employed to optimize chromosome doubling for tetraploid production as they are typically used by vegetative propagation. However, they have the popularity as ornamentals globally due to their horticultural aspects. Tetraploid induction was optimized by in vitro somatic chromosome doubling on H. coronarium and H. gardnerianum […]

  • In vitro culture in combination with aeroponics is an efficient means of mass propagation of Sarcostemma acidum: a rare medicinal plant of Indian arid zone
    on June 1, 2022 at 12:00 am

    Abstract In vitro culture in combination with aeroponics is observed to be an efficient means for mass propagation of Sarcostemma acidum in the present investigation. S. acidum is a rare leafless xerophytic shrub of the family Apocynaceae, commonly known as Som-lata or khir-kheep, a source of medicines and the religious drink “Somaras.” Aeroponic technique was used for the induction of adventitious roots from stem cuttings of field-grown plants for the development of quality plants in the greenhouse as a source of explants for micropropagation. Among various concentrations of IBA, NAA and NOA used for induction of adventitious rooting from stem cuttings 3.0 g L−1 NAA were found to be most suitable with average 90% induction response and 14 adventitious roots. Surface-sterilized nodal shoot segments of 6 to 7 cm length were inoculated on basic and modified Murashige and Skoog’s medium containing 3% (w/v) sucrose, 0.8% (w/v) agar, and various concentrations […]

  • In vitro propagation, lactucin quantification, and antibacterial activity of Indian lettuce (Lactuca indica L.)
    on June 1, 2022 at 12:00 am

    Abstract  Lactuca indica L. (Asteraceae), a wild lettuce, is used as a vegetable and in traditional medicine. This study aims to establish in vitro propagation protocol and evaluate lactucin and antibacterial property from in vitro and natural plant tissues. Leaf blades and petioles were cultured in vitro on Murashige and Skoog (MS) medium supplemented with various concentrations of 6-benzylaminopurine (BAP) and 1.2 mg L–1 indoleacetic acid (IAA). Leaf petioles and a lower BAP concentration (0.5 mg L–1) were optimal for direct shoot induction, while the leaf blade and a higher BAP (4 mg L–1) concentration performed best for callus induction. When the callus was subcultured, 98.7% of samples regenerated plants on MS medium supplemented with 1.5 mg L–1 BAP and 0.5 mg L–1 IAA. MS medium containing 1 mg L–1 IAA was best for in vitro rooting. A high-performance liquid chromatography analysis of the in vitro samples revealed a higher amount of lactucin […]

  • Shoot organogenesis and plant regeneration in Cordia subcordata Lam
    on June 1, 2022 at 12:00 am

    Abstract Cordia subcordata Lam. (Boraginaceae) is a woody tree that grows in coastal wasteland and mangrove edges. Its wood is used for woodcraft, and the tree, which is used in landscaping, also has medicinal and ecological value (e.g., coastal protection). In this study, a shoot proliferation and regeneration system was established for this plant. Callus was induced from terminal young leaf explants on Murashige and Skoog (MS) medium with 1.0 mg L–1 thidiazuron or 1.0 mg L–1 2,4-dichlorophenoxyacetic acid, or optimally with 0.5 or 1.0 mg L–1 of both. Callus differentiated from an average of 35.4% of explants into 3 to 5 adventitious shoots (indirect shoot organogenesis) within 45 d. Optimal shoot proliferation medium was MS medium with 0.8 mg L–1 6-benzyladenine (BA). A higher concentration of BA (1.5 mg L–1) in the presence of α-naphthaleneacetic acid (NAA) enhanced callus induction from the base of shoots and decreased shoot […]

  • NanoAOX—a novel nanoparticle and antioxidant mixture enhances the growth of plants in vitro and in vivo
    on June 1, 2022 at 12:00 am

    Abstract The global food crisis is an issue affecting 1 billion people worldwide—it is critical that a solution be developed to provide the world’s citizens with alternative sources of food that are less energy-intensive. Both nanoparticles, molecules < 100 nm used in drug delivery, and antioxidants, molecules that inhibit oxidation reactions, have been individually studied to enhance plant growth; however, the combined effect has not been investigated. Using hydroponic, tissue culture, and hydrogel experiments, this project investigates whether a combined nanoparticle-antioxidant treatment of carbon nanoparticles and ascorbic acid (CNP-AOX) has a synergistic effect in enhancing plant growth. When tested using hydroponic experiments, NP-AOX showed significant increases in chlorophyll levels, media consumption, and plant weight compared to the other treatments. Tissue culture experiments showed that nanoparticles resulted in long roots, antioxidants resulted […]

  • Effective in vitro culture using dormant bud of nodal sections from a mature Acacia tree
    on June 1, 2022 at 12:00 am

    Abstract The micropropagation system described is a commercially viable method for asexual propagation of a mature ‘elite’ tree of Acacia confusa. The main stem of a mature tree was hardly pruned to induce new shoots for the explants. This study showed that PPM™ (Plant Preservative Mixture) was an effective agent for controlling the contamination of nodal explants and was added to the initial culture medium. This medium consisted of Murashige and Skoog (MS) basal salts and vitamins, supplemented with 4.0 mg L−1 BA, 0.1 mg L−1 IBA, 10 mg L−1 GA, 12 mg L−1 AS, 3.2 g L−1 PVP, and 2.0 mL L−1 PPM™, and was adequate for the establishment of in vitro mother stock for proliferation. The excellent growth response of nodal explants, obtained from May to July in 2019 to 2020, had shoot formation rate ranging from 15.8 to 18.5%. The optimal shoot multiplication was derived on MS medium supplemented with 1.0 mg L−1 BA, 0.5 mg L−1 TDZ, […]

  • Development of a high-frequency adventitious shoot regeneration using cotyledon explants of an important oilseed crop Sesamum indicum L.
    on June 1, 2022 at 12:00 am

    Abstract Sesame (Sesamum indicum L.; 2n = 26) is an ancient oil-yielding crop known for its valuable nutrients such as essential fatty acids, proteins, and medicinally important lignans. Recalcitrant nature of sesame limits the use of modern biotechnology for its genetic improvement. For the development of an efficient in vitro regeneration protocol, the cotyledon (de-embryonated) explants were used for the development of adventitious shoots. In the present study, Murashige and Skoog (MS) medium with 25 μM BAP (6-benzylaminopurine) and 2 μM IAA (indole-3-acetic acid) showed high regeneration frequency in both GT-3 (33.3 ± 1.1%) and GT-4 (30 ± 0.0%) varieties within 6 wk of culture. Addition of 20 μM thidiazuron (TDZ) showed further improvement in regeneration frequency in both GT-3 (63.3 ± 1.1%) and GT-4 (26.7 ± 1.1%) varieties. In another experiment, 0-d-old cotyledons inoculated on MS medium supplemented with 4 μM abscisic acid […]

  • Agrobacterium-mediated transformation of purple raspberry (Rubus occidentalis × R. idaeus) with the PtFIT (FER-like iron deficiency–induced transcription factor 1) gene
    on June 1, 2022 at 12:00 am

    Abstract An Agrobacterium-mediated transformation protocol for the purple raspberry (R. occidentalis × R. idaeus) ‘Amethyst’ was developed. Using the system, the FER-like iron deficiency–induced transcription factor 1 gene cloned from Populus tremula (PtFIT) was expressed in transgenic raspberry plants. Effects of four inoculum densities and two co-cultivation times on ‘Amethyst’ transformation were tested in two separate experiments. Results showed that an average transformation frequency of 3.9% was achieved under the conditions of 25 mg L−1 kanamycin selection, 3-d co-cultivation, and OD600 0.3 to 0.55 inoculum density. A total of 12 PtFIT-transgenic lines of ‘Amethyst’ were verified using polymerase chain reaction (PCR) analysis. Expression of the PtFIT gene in transgenic lines was evaluated under the iron deficiency or sufficiency condition using the real-time quantitative PCR (RT-PCR); however, the expression showed an […]

  • Coat protein–mediated resistance to cucumber mosaic virus subgroup IB in black pepper (Piper nigrum L.)
    on June 1, 2022 at 12:00 am

    Abstract Cucumber mosaic virus (CMV) infects a large number of plant species including Piper nigrum L. and related species. As natural resistance to CMV is absent in Piper spp., the study was undertaken to produce transgenic P. nigrum plants harboring the complete coat protein (CP) gene of CMV via Agrobacterium-mediated transformation and their evaluation for resistance against the virus. Among one hundred and nine hardened transformed plantlets, eight revealed the presence of the transgene in PCR. The production of transcript in these plants was assessed by reverse transcription-polymerase chain reaction (RT-PCR) and buildup of CMV CP by direct antigen-coated enzyme-linked immunosorbent assay (DAC-ELISA). Screening of all eight transgenic lines against CMV through cleft grafting revealed that all lines except one were symptomless or showed mild or moderate symptoms. The transgenic line with the highest resistance was vegetatively propagated and integration of […]

  • Micropropagation, phytochemical content and antioxidant activity of gamma-irradiated blackberry (Rubus fruticosus L.) plantlets
    on June 1, 2022 at 12:00 am

    Abstract In vitro methods were used to study the consequences of gamma irradiation on morphological and biochemical changes in blackberry plantlets. Gamma-irradiated (20, 40, and 60 Gy) plantlets were subcultured using Murashige and Skoog (MS) media. Each irradiation treatment dose consisted of ten jars containing three shoots per jar. Methanolic extracts were used for determination of total phenolic content, phenolic acids, flavonoid contents, antioxidant activity using DPPH, and ascorbic acid. The rise in γ-irradiation dose (40 Gy) significantly increased percent rooting. Raising the dose level of γ-irradiation to 40 Gy in the first, second and third subcultures of Rubus fruticosus plantlets resulted in significant enhancement of vegetative traits. In plantlets exposed to 40 Gy γ-irradiation, a significant increase was observed in chlorophyll a, b and carotenoid levels. Meanwhile, chlorophyll and carotenoid content decreased when the irradiation dose level […]

  • Elicitation enhanced the production of bioactive compound and biomass accumulation in callus cultures of Glycyrrhiza glabra L.
    on June 1, 2022 at 12:00 am

    Abstract Licorice (Glycyrrhiza glabra L.), Fabaceae, has a wide range of medicinal properties due to the metabolites found in the plant tissues. Callus cultures from Glycyrrhiza glabra were previously initiated in vitro using leaf as an explant and the current study was designed to examine the possible role of various elicitors (biotin, adenine sulphate, salicylic acid, putrescine, spermine, or spermidine) at different concentrations (25, 50, 75, and 100 mg L−1) on the stimulation of biomass and their effects on the content of different metabolites such as total carbohydrates, protein, proline, phenol, alkaloids, flavonoids, and glycyrrhizin. Elicitation with different concentrations of elicitors increased the biomass and metabolite content in callus culture of G. glabra at different rates. The optimum concentration of adenine sulphate for maximum biomass accumulation (16.79 g flask−1) was found to be at 50 mg L−1 after incubation for 20 d. Adenine […]

  • Phytochemical and anti-oxidant evaluation of in vitro and in vivo propagated plants of Curculigo orchioides
    on June 1, 2022 at 12:00 am

    Abstract Curculigo orchioides (Kali Musli) is a medicinal plant commonly used in the Indian and Chinese traditional medicine system to treat jaundice and asthma. It is also used as an aphrodisiac in several countries. C. orchioides is treated as an essential resource in many pharmaceutical industries as a source of bioactive compounds. Indiscriminate use, depletion of its natural habitat, and its seasonal availability have resulted in its rarity. As per the IUCN status, Curculigo orchioides is now listed as an endangered plant. The present micropropagation study resolves its seasonal availability by ensuring a continuous supply of plants. In vitro somatic embryos were induced on the leaf explants of C. orchioides in one-fourth–strength Murashige and Skoog (MS) media. The embryos developed into whole plants in 8 wk. The in vitro plants produced were evaluated for their phytochemical evaluation and anti-oxidant activity along with their wild counterparts as […]

  • Adventitious bud regeneration and Agrobacterium tumefaciens-mediated genetic transformation of Eucalyptus urophylla × E. tereticornis interspecific hybrid
    on June 1, 2022 at 12:00 am

    Abstract A high-efficiency regeneration and genetic transformation system is indispensable for generating desirable traits in important trees such as Eucalyptus. However, lower regeneration efficiency is common for most varieties because of the recalcitrance of this genus. Here, a stable and highly efficient in vitro organogenesis protocol and Agrobacterium-mediated genetic transformation system of Eucalyptus were developed, and transgenic plants were obtained. In this protocol, the preferred explants were the top and middle stem internodes from in vitro micro-shoots of the E. urophylla × E. tereticornis hybrid. Modified Woody Plant Medium (mWPM) containing 0.025 mg·L-1 thidiazuron (TDZ) and 0.10 mg·L-1 indole-3-butyric acid (IBA) was used to induce multiple adventitious buds that allowed 85.6% shoot formation. The binary vector pBI121 carrying the neomycin phosphotransferase II (nptII) and β-glucuronidase (uidA) genes was applied for transformation. The […]

  • Callus induction and multiple shoot proliferation from nodal explants of Mansonia altissima: confirmation of genetic stability using ISSR and RAPD markers
    on June 1, 2022 at 12:00 am

    Abstract This study aimed to investigate indirect micropropagation of Mansonia altissima using nodal explant and confirmation of genetic stability of the plantlets in order to establish the protocol for callus induction, mass propagation, and true to type clones. Murashige and Skoog (MS) medium supplemented with 1.0, 2.0, 4.0, 8.0, and 10.0 μM IAA (indole-3-acetic acid) or 1.0, 2.0, 4.0, 8.0, and 10.0 μM IBA (indole-3-butyric acid) and 50.0 mg·L−1 l-ascorbic acid was used for callus induction. The callus was subcultured on Murashige and Skoog (MS) medium containing BAP (6-benzylaminopurine) (μM) or TDZ (thidiazuron) (μM) and KIN (kinetin) (μM). The induced shoots were cultured on different strength MS medium, activated charcoal, and indole-3-butyric acid (μM) for root induction. The plantlets were acclimatized to the environment using four different types of potting media. Polymerase chain reaction amplification of the wild and indirect micropropagated […]

  • In vitro assessment of strawberry (Fragaria × ananassa Duch.) plant responses to water shortage stress under nano-iron application
    on June 1, 2022 at 12:00 am

    Abstract Strawberry is a susceptible plant to water stress, and its growth is severely declined under water shortage. Therefore, the aim of this study was to assess the impacts of iron, as a micro-nutrient, in the form of a nano-oxide on the tolerance of strawberry plants to water stress conditions under in vitro culture. Three levels of PEG-induced water stress (0, 5%, and 7% w v−1 PEG 6000) and three levels of iron nano-particles (0, 1, and 1.2 mg L−1) were supplemented to Murashige and Skoog (MS) culture medium. The results demonstrated that PEG-induced water stress negatively affected shoot fresh and dry weight, root fresh and dry weight, and physiological traits such as relative water content, membrane stability index, and pigment content. The biochemical traits were also affected by PEG-induced water stress; consequently, antioxidant enzyme activity, leaf proline content, total soluble sugar, and malondialdehyde (MDA) and H2O2 content were increased. The […]

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