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.


  • In vitro desiccation tolerance of the epiphytic Ghost Orchid, Dendrophylax lindenii (Lindl.) Benth x. Rolfe
    on December 7, 2018 at 12:00 am

    Abstract The Ghost Orchid, Dendrophylax lindenii (Lindl.) Benth x. Rolfe, is a rare and endangered epiphytic orchid native to south Florida and Cuba. The orchid is considered difficult to propagate under greenhouse conditions, requiring high humidity and low air movement. In contrast, the orchid’s native habitat seasonally dries out with decreased precipitation and humidity. This suggests some level of desiccation tolerance. Ghost Orchid plants were assessed for potential desiccation tolerance and ability to recover from desiccation stress under in vitro conditions. In vitro-derived plants were placed into sterile baby food jars and transferred to chambers maintained at 10% relative humidity, which is extremely low compared to relative humidity levels (40–100%) recorded under natural field conditions. Plants were removed every week for 4 wk and recovered on P723 medium supplemented with banana powder for 4 wk. Data were collected at the […]

  • Droplet-vitrification cryopreservation of in vitro -grown shoot tips of grapevine ( Vitis spp.)
    on December 1, 2018 at 12:00 am

    Abstract An efficient and broad-spectrum protocol for cryopreservation of Vitis spp. shoot tips by droplet-vitrification is reported. Shoot tips (1.0 mm) containing 5–6 leaf primordia (LPs) were precultured for 3 d with a preculture medium containing 0.3 M sucrose, 0.16 μM glutathione, and 0.14 μM ascorbic acid. Precultured shoot tips were treated for 20 min at 24°C with a loading solution composed of 2 M glycerol and 0.4 M sucrose, followed by exposure at 0°C to half-strength plant vitrification solution 2 (PVS2) for 30 min, and then full-strength PVS2 for 50 min. Dehydrated shoot tips were transferred into 2.5-μL PVS2 carried on aluminum foil, prior to a direct immersion in liquid nitrogen. With this method, an average shoot regrowth level of 50.5% was obtained from cryopreserved shoot tips in six V. vinifera genotypes (three wine cultivars, two table cultivars, and one rootstock) and two V. […]

  • Micropropagation of Glossonema varians (Stocks) Benth. ex Hook.f.—a rare Asclepiadeae of Indian Thar Desert
    on December 1, 2018 at 12:00 am

    Abstract In the present study, an in vitro regeneration protocol for Glossonema varians (Stocks) Benth. ex Hook.f. of family Asclepiadaceae was optimized. Cotyledonary nodes of in vitro cultured seedlings were used as explants for activation of axillary shoot buds. Axillary shoot buds were initially activated on 0.1 mg L−1 6-benzylaminopurine (BAP) and then multiplied on 0.05 mg L−1 BAP. Shoots were rooted in vitro on 1/4 strength Murashige and Skoog medium containing 0.1 mg L−1 2-naphthoxyacetic acid and 100 mg L−1 activated charcoal. The cultures were maintained in a 12 h photoperiod at 40–50 μmol m−2 s−1 spectral flux photon, 25–30 ± 2°C, and 60% relative humidity (RH). Up to 80% of in vitro regenerated plantlets were acclimatized on soilrite in cotton-plugged culture tubes in the greenhouse. This protocol can be a useful method […]

  • In vitro growth profile and comparative leaf anatomy of the C 3 –C 4 intermediate plant Mollugo nudicaulis Lam.
    on December 1, 2018 at 12:00 am

    Abstract Mollugo nudicaulis Lam., commonly known as John’s folly or naked-stem carpetweed, is an ephemeral species of tropical regions. The plant is ideal to study the eco-physiological adaptations of C3–C4 intermediate plants. In the present report, in vitro growth profiling of the plant and comparative leaf anatomy under in vitro and ex vitro conditions were studied. In vitro propagation of the plant was carried out on Murashige and Skoog (MS) basal medium augmented with additives and solidified with 0.8% (w/v) agar-agar or 0.16% (w/v) Phytagel™. The concentration of plant growth regulators (PGRs) in the basal medium was optimized for callus induction, callus proliferation, shoot regeneration, and in vitro rooting. The optimum callus induction was obtained from M. nudicaulis seedling hypocotyls. The highest regeneration induction of about 88% or nearly 41 shoots with about 142 leaves per culture vessel was observed from friable callus on MS […]

  • In vitro preservation and micropropagation of Oreocharis mileense (W.T. Wang) M. Möller & A. Weber (Gesneriaceae) through shoot organogenesis
    on December 1, 2018 at 12:00 am

    Abstract Oreocharis mileense (W.T. Wang) M. Möller & A. Weber is endemic to China and was considered to be extinct because it had not been seen in the wild since the first collection in 1906. In 2006, the species was rediscovered in Shilin County, Yunnan Province. Oreocharis mileense was considered critically endangered for its narrow geographic range and extremely small population. An efficient method to preserve plant germplasm by in vitro culturing of O. mileense has not been reported. In this study, an orthogonal array with three factors (6-benzyladenine, BA; α-naphthaleneacetic acid, NAA; and sucrose), at four levels was performed, and shoot induction as well as shoot proliferation were recorded. The results were analyzed to determine the most significant components and the optimum combination for micropropagation of O. mileense. The results showed that: (1) organogenesis was easily induced by different combinations of plant-growth regulators […]

  • Efficient callus-mediated regeneration and in vitro root tuberization in Trichosanthes kirilowii Maxim., a medicinal plant
    on December 1, 2018 at 12:00 am

    Abstract Trichosanthes kirilowii Maxim. is a climbing herb with considerable medicinal value. In this study, efficient protocols for callus-mediated regeneration and in vitro tuberization of this plant were developed. Sterilized stem and leaf tissues were cultured on Murashige and Skoog (MS) medium with plant growth regulators (PGRs), and additives that promoted callus induction and regeneration. Both stem and leaf tissues showed the best response (100%) for callus initiation on MS medium supplemented with 4.5-μM 2,4-dichlorophenoxyacetic acid (2,4-D). Efficient shoot organogenesis was obtained by exposing the callus tissue to 4.6-μM kinetin, 2.2-μM 6-benzylaminopurine, and 2.7-μM 1-naphthylacetic acid (NAA) along with 12.6-μM copper sulfate, which yielded a shoot regeneration rate of 85.5% and 28 shoots derived from each callus. In vitro shoots were best rooted on half-strength (1/2) MS medium with 2.7-μM NAA. Tuberous roots were efficiently […]

  • Validation of reference genes for gene expression analysis of response to anthocyanin induction in cell cultures of Vitis davidii (Rom. Caill.) Foëx
    on December 1, 2018 at 12:00 am

    Abstract Real-time quantitative polymerase chain reaction (RT-qPCR) is an effective method for detecting changes of gene expression in plant cell metabolic regulation. A set of 15 reference gene candidates were selected for the present study of anthocyanin biosynthesis regulation, and stability. The suitability of their expression was evaluated in eight different experimental treatments in spine grape (Vitis davidii [Rom. Caill.] Foëx.) cell cultures. The results indicated that SAND family protein (SAND) and V-type proton ATPase subunit G (VAG) were the most stable reference genes for culture duration, tubulin alpha-3/alpha-5 chain (α-tubulin) and tubulin beta-1 chain (β-tubulin) for illumination conditions, ubiquitin-conjugating enzyme E2-17 kDa (UBQ) and VAG for UVB treatment, VAG and 60S ribosomal protein L18-2 (60SRP) for temperature treatment, AP47, clathrin adaptor complex subunit mu (AP-2) and 60SRP for cinnamic acid treatment, […]

  • Micropropagation of virus-free plants of Saudi fig ( Ficus carica L . ) and their identification through enzyme-linked immunosorbent assay methods
    on December 1, 2018 at 12:00 am

    Abstract Viral infection is one of the most serious biotic stresses, which disturbs the growth and productivity of many horticultural crops, including that of fig (Ficus carica L.). The production of plants free of viruses, such as fig mosaic virus (FMV), has become a priority in many plant breeding programs. In this study, leaves from plants of two fig cultivars, Kodato and Dattora, infected with FMV were collected from both Mecca and Al-Taif, Saudi Arabia. Transmission electron microscopy of ultrathin leaf sections showed double membrane bodies, characteristic of FMV particles, only in the mesophyll cells of infected samples. Protein analysis using sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the presence of a protein band with a molecular weight of 35 kDa, which corresponded to the viral coat protein; and FMV was confirmed by Western blot and enzyme-linked immunosorbent assay (ELISA) tests. To obtain virus-free plants, apical shoot […]

  • Improved and synchronized maturation of Norway spruce ( Picea abies (L.) H.Karst.) somatic embryos in temporary immersion bioreactors
    on December 1, 2018 at 12:00 am

    Abstract Somatic embryogenesis offers many benefits for clonal propagation in large-scale plant production of conifers. A key rate-limiting step is the conversion from early-stage somatic embryos in pro-embryogenic masses (PEMs) to the maturation stage. Immature embryos in PEMs are present at different developmental stages, where some are unable to respond to the maturation treatment, thus limiting yields of mature embryos. Synchronization of early somatic embryo development in PEMs could greatly improve subsequent yields of mature embryos. A temporary immersion bioreactor designed for Norway spruce (Picea abies (L.) H.Karst.) was used in this study. Through a specific system for dispersion, connected tissue of PEMs, composed of immature embryos grown in liquid medium in the temporary immersion bioreactors or on solid medium as a control, was dispersed and redistributed in a more uniform spatial arrangement. It was demonstrated that development of mature embryos […]

  • Quercetin and silver nitrate modulate organogenesis in Carissa carandas (L.)
    on December 1, 2018 at 12:00 am

    Abstract An in vitro organogenesis protocol for Carissa carandas L. was developed using an auxin transport inhibitor (quercetin) and silver nitrate (AgNO3), an inhibitor of ethylene action, in association with cytokinins in the culture medium. This protocol produced the maximum number of shoots from aseptic seedling-derived shoot apex explants of C. carandas. The highest rate of shoot multiplication was recorded on MS medium containing 2.0 mg L−1 6-benzylaminopurine; 0.5 mg L−1 kinetin, and 0.75 mg L−1 quercetin at after 4 wk of culture. Similar results were obtained when MS medium fortified with 2.0 mg L−1 BAP, 0.5 mg L−1 kinetin, and 1.5 mg L−1 AgNO3 was used. However, successful rooting was achieved on quarter strength MS medium with 0.5 mg L−1 indole-3-acetic acid. In this study, an inhibitor of auxin transport and ethylene action maximized shoot […]

  • Improved Agrobacterium tumefaciens- mediated transformation of soybean [ Glycine max (L.) Merr.] following optimization of culture conditions and mechanical techniques
    on December 1, 2018 at 12:00 am

    Abstract In the present study, Agrobacterium tumefaciens-mediated transformation of Glycine max (L.) Merr. (soybean) cv. DS-9712 using half-seed explants was optimized for eight different parameters, including seed imbibition, medium pH, infection mode (sonication and vacuum infiltration), co-cultivation conditions, concentrations of supplementary compounds, and selection. Using this improved protocol, maximum transformation of 14% and regeneration efficiencies of 45% were achieved by using explants prepared from mature seeds imbibed for 36 h, infected with A. tumefaciens strain EHA105 at an optical density (OD600) of 0.8, suspended in pH 5.4 medium containing 0.2 mM acetosyringone and 450 mg L−1 L-cysteine, followed by sonication for 10 s, vacuum infiltration for 2 min, and co-cultivated for 3 d on 35 mg L−1 kanamycin-containing medium. Independent transgenic lines were confirmed to be transgenic after […]

  • A filter paper-based liquid culture system for citrus shoot organogenesis—a mixture-amount plant growth regulator experiment
    on December 1, 2018 at 12:00 am

    Abstract This study determined the effects of a static liquid culture system on shoot regeneration from citrus epicotyl explants. A mixture-amount experiment was used to determine the effects of zeatin riboside (ZR), 6-benzylaminopurine (BA), and indole-3-acetic acid (IAA) on two citrus types—citrange (Citrus sinensis ‘Washington’ L. Osbeck. × Poncirus trifoliata L. Raf var. Carrizo) and sweet orange (Citrus sinensis L. Osbeck var. ‘Ridge Pineapple’). A liquid culture system comprising a Petri dish, cellulose filter paper, and liquid culture medium was used. Shoot regeneration experiments were conducted over 6 wk that included 2 wk in the dark followed by 4 wk in the light. Three responses were measured: (1) number of explants forming buds and/or shoots, (2) number of explants with shoots > 2 mm, and (3) overall explant and shoot quality. The effects of paper disc number, liquid medium volume, and […]

  • Responses of Arabica coffee ( Coffea arabica L. var. Catuaí) cell suspensions to chemically induced mutagenesis and salinity stress under in vitro culture conditions
    on December 1, 2018 at 12:00 am

    Abstract Crop improvement of Coffea arabica L. (coffee) via mutagenesis could accelerate breeding programs; thus, the present study aimed to develop an in vitro protocol using the chemical mutagens sodium azide (NaN3) and ethyl methanesulfonate (EMS) on embryogenic cell suspensions of Arabica coffee variety Catuaí and, subsequently, to evaluate the responses of the resulting mutagenized tissues to salinity stress. Embryogenic suspension cultures were incubated with 0.0, 2.5, 5.0, or 10.0 mM NaN3 or 0.0, 185.2, 370.5, or 741.0 mM EMS. As the concentration of NaN3 or EMS increased, the survival of embryogenic suspension cultures decreased compared to controls. The median lethal dose (LD50) for NaN3 was 5 mM for 15 min and for EMS it was 185.2 mM for 120 min. Embryogenic suspension cultures treated with NaN3 or EMS were cultured on selective medium supplemented with 0, 50, 100, 150, 250, or 300 mM NaCl showed that 50 mM […]

  • Cryopreservation of an endangered Hladnikia pastinacifolia Rchb. by shoot tip encapsulation-dehydration and encapsulation-vitrification
    on December 1, 2018 at 12:00 am

    Abstract The objective of the present study was the cryopreservation of monotypic endemic Hladnikia pastinacifolia Rchb. shoot tips from an in vitro culture, via encapsulation-dehydration (ED) or encapsulation-vitrification (EV). For all tested genotypes, the highest rates of shoot regrowth and multiplication were obtained after overnight preculture in 0.4 M sucrose, encapsulation in Murashige and Skoog (MS) medium with 0.4 M sucrose and 1 M glycerol, followed by polymerization in 3% (w/v) Na-alginate in MS with 0.4 M sucrose. Optimal osmoprotection was achieved for ED with 0.4 M sucrose plus 1 M glycerol and for EV with 0.4 M sucrose plus 2 M glycerol. The best dehydration time for ED was 150 min in a desiccation chamber with silica gel, and the best vitrification time for EV was 85 min in plant vitrification solution 2 (PVS2). For ED, dehydration for 150 min resulted in explant water content of 22%. When the […]

  • An efficient protocol for in vitro regeneration and conservation of Shirui lily ( Lilium mackliniae Sealy): a lab-to-land approach to save the rare endangered Asiatic lily species
    on December 1, 2018 at 12:00 am

    Abstract An efficient protocol for direct and indirect shoot regeneration and proliferation from bulb scales of Shirui lily (Lilium mackliniae Sealy), an endangered Asiatic lily species endemic to the Shirui hill peak, Manipur, India, has been developed. Bulb scales were isolated from mature bulbs and cultured on Murashige and Skoog (MS) basal medium supplemented with different concentrations of 6-benzylaminopurine (BAP), kinetin (KIN), or thidiazuron (TDZ). For direct shoot regeneration from bulb scale explants, 0.5 mg L−1 BAP yielded the highest shoot induction (3.5 shoots per scale; a 96.7% response). For indirect de novo organogenesis, optimum callus induction was achieved with 2.0 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D), and shoot organogenesis was higher (16.2) when subcultured onto 0.5 mg L−1 BAP medium. Multiple shoot regeneration and pseudo-bulb formation protocols were assessed; the highest shoot […]

  • In vitro micrografting using three diverse indigenous rootstocks for the production of Citrus tristeza virus- free plants of Khasi mandarin
    on November 8, 2018 at 12:00 am

    Abstract The present work reports the prospective applicability of three indigenous rootstocks belonging to different species viz. Nemutenga, Tayum and Tasi cultivars of north eastern Himalayan region for producing Citrus tristeza virus (CTV; a viral species of the Closterovirus genus) free quality planting materials of Khasi mandarin (Citrus reticulata Blanco) through micrografting or shoot-tip grafting (STG). The development of disease-free plants through STG is essential as CTV is in vivo transmissible through infected bud sticks. The technique of STG along with the diverse culture parameters like effect of sucrose, plant growth regulators, pre-treatment of scion and stock, effect of scion size after thermotherapy and grafting method was analysed. Among the three indigenous tested rootstocks, Nemutenga was found superior showing maximum STG success of 58.5%. STGs using scion size ranging from 0.3 to 0.5 mm, survival was up to 42.0% and completely virus-free. […]

  • Cell suspension culture of Rhizoma zedoariae in a two-stage perfusion bioreactor system for β-elemene production
    on November 5, 2018 at 12:00 am

    Abstract We presented a two-stage combined bioreactor system consisting of a stir-tank and an airlift column, and challenged with Rhizoma zedoariae cell suspensions for β-elemene production. Two-stage culture was initiated when the cell concentration in both vessels was maintained at an appropriate density. The cells were proliferated in stirred-tank with the maximal growth rate of 0.17 d−1 to present enough cells for β-elemene synthesis. In the airlift column, continuous cell separation from culture medium was achieved by using a cell retention device based on centrifugal and gravity settling when the system was performed in perfusion mode. The results indicated that additives can efficiently promote the accumulation of β-elemene in R. zedoariae cells. In addition, the β-elemene content showed higher levels in cell lines of overexpressing 3-hydroxy-3-methylglutaryl coenzyme-A reductase, Farnesyldi phosphate synthase, and ST02C genes. […]

  • Data mining study of hormone biosynthesis gene expression reveals new aspects of somatic embryogenesis regulation
    on October 30, 2018 at 12:00 am

    Abstract Although the application of exogenous hormones and the status of endogenous hormones play a crucial factor in somatic embryogenesis (SE), our understanding of the status of endogenous hormones and the molecular framework for hormone biosynthesis during SE are still elusive. With a data mining approach, we analyzed public RNA-seq data of SE from two model plants, Arabidopsis thaliana and Oryza sativa var. Japonica, which showed that several plant hormone biosynthesis genes had distinct patterns during SE. The expressions of auxin, cytokinin, gibberellin, and ethylene biosynthesis genes were induced during SE in A. thaliana, while only jasmonic acid showed an expression increase during somatic embryogenesis in both plants. Furthermore, by analyzing public RNA sequencing and DNA affinity purification sequencing (DAP-seq), we revealed some possibilities of the role of ethylene during SE through the activity of EIN3, by increasing the expression level of major […]

  • Cloning and characterization of Somatic Embryogenesis Receptor Kinase I ( EgSERK I ) and its association with callus initiation in oil palm
    on October 9, 2018 at 12:00 am

    Abstract The somatic embryogenesis receptor kinase (SERK) gene has been extensively studied in many plant species due to its role in conferring embryogenic competence to somatic cells. The oil palm (Elaeis guineensis Jacq.) full-length SERK I (EgSERK I) cDNA was first isolated from cell suspension culture using RACE-PCR. Total length of EgSERK I cDNA was 2378 bp in length with a 5’UTR region (358 bp) longer than 3’UTR region (130 bp) and the ORF was 1890 bp (629aa). The deduced amino acid sequence of EgSERK I contained protein domains commonly present in reported SERK proteins, including the hallmark proline-rich region and C-terminal domains. EgSERK I was most highly expressed in leaf explants and also detected in all tested tissues, including vegetative tissues, reproductive tissues, embryogenic tissues, and non-embryogenic tissues, suggesting that it may have a broad role in plant growth and development. Expression of EgSERK I in […]

  • Application of wide-spectrum light-emitting diodes in micropropagation of popular ornamental plant species: a study on plant quality and cost reduction
    on October 9, 2018 at 12:00 am

    Abstract In the present study, the applicability of four wide-spectrum light-emitting diodes (LEDs) emitting warm light (AP67, AP673L, G2, and NS1) was determined for the micropropagation of five popular ornamental plant species: Chrysanthemum × grandiflorum, Gerbera jamesonii, Heuchera × hybrida, Ficus benjamina, and Lamprocapnos spectabilis. Plantlets were grown in a growth room with a 16-h photoperiod. The photosynthetic photon flux density was set at 62–65 μmol m−2 s−1. The composition of the media and subculture timing were adjusted to the needs of each species tested. The results were compared to the cool daylight-emitting fluorescent (FL) control (TLD 36W/54). In most of the species studied (except for F. benjamina), the highest propagation ratios, or ratios similar to the FL control, were observed under the red- and far-red-abundant G2 LEDs. NS1 spectrum (with the highest proportion of blue and green light) was […]

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