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PubMed2002

[Study on stem-tip tissue culture of the traditional Chinese medicine Chrysanthemum morifolium].

OBJECTIVE: To set up the optimums for the stem-tip tissue culture of Chrysanthemum morifolium cultivated in Anhui Province. METHOD: Small sections (about 0.5 mm in length) from the stem-tips were isolated and inoculated with different media, and induced to form the whole plantlet formation. RESULT AND CONCLUSION: The MS medium added with 6-BA 2 mg.L-1 + NAA 0.2 mg.L-1 was the optimum medium for the bud sprouting and the inducing rate was over 80% after 40 d cultivation on this modified medium. The MS medium supplemented with 6-BA 2 mg.L-1 and NAA 0.5 mg.L-1 was the optimum medium for the multiplication of the adventitious buds in which the bud multiplication was about 4-7 times higher after 25-30 d cultivation. The plantlet could root well on the MS medium with NAA 0.5 mg.L-1.

Chrysanthemum↗

PubMed2003

Efficient regeneration of sorghum, Sorghum bicolor (L.) Moench, from shoot-tip explant.

Novel protocols for production of multiple shoot-tip clumps and somatic embryos of Sorghum bicolor (L.) Moench were developed with long-term goal of crop improvement through genetic transformation. Multiple shoot-tip clumps were developed in vitro from shoot-tip explant of one-week old seedling, cultured on MS medium containing only BA (0.5, 1 or 2 mg/l) or both BA (1 or 2 mg/l) and 2,4-D (0.5 mg/l) with bi-weekly subculture. Somatic embryos were directly produced on the enlarged dome shaped growing structures that developed from the shoot-tips of one-week old seedling explants (without any callus formation) when cultured on MS medium supplemented with both 2,4-D (0.5 mg/l) and BA (0.5 mg/l). However, the supplementation of MS medium with only 2,4-D (0.5 mg/l) induced compact callus without any plantlet regeneration. Each multiple shoot-clump was capable of regenerating more than 80 shoots via an intensive differentiation of both axillary and adventitious shoot buds, the somatic embryos were capable of 90% germination, plant conversion and regeneration. The regenerated shoots could be efficiently rooted on MS medium containing indole-3-butyric acid (IBA 1 mg/l). The plants were successfully transplanted to glasshouse and grown to maturity with a survival rate of 98%. Morphogenetic response of the explants was found to be genotypically independent.

Plant Shoots↗

PubMed2006

Factors affecting in vitro plant regeneration of the critically endangered Mediterranean knapweed (Centaurea tchihatcheffii Fisch et. Mey).

Habitat destruction has resulted in the extinction of many plant species from the earth, and many more face extinction. Likely, the annual endemic Mediterranean knapweed (Centaurea tchihatcheffii) growing in the Golbasi district of Ankara, Turkey is facing extinction and needs urgent conservation. Plant tissue culture, a potentially useful technique for ex situ multiplication, was used for the restoration of this ill-fated plant through seed germination, micropropagation from stem nodes, and adventitious shoot regeneration from immature zygotic embryos. The seeds were highly dormant and very difficult to germinate. No results were obtained from the micropropagation of stem nodes. However, immature zygotic embryos showed the highest adventitious shoot regeneration on Murashige and Skoog (MS) medium, containing 1 mg l(-1) kinetin and 0.25 mg l(-1) NAA. Regenerated shoots were best rooted on MS medium containing 1 mg l(-1) IBA and transferred to the greenhouse for flowering and seed set. As such, the present work is the first record of in vitro propagation of critically endangered C. tchihatcheffii, using immature zygotic embryos, and is a step forward towards conservation of this indigenous species.

Centaurea↗

PubMed2005

Factors influencing axillary shoot proliferation and adventitious budding in cedar.

We developed procedures for in vitro cloning of Cedrus atlantica Manetti and C. libani A. Rich explants from juvenile and mature plants. Explant size was one determinant of the frequency of axillary bud break in both species. Shoot tips and nodal explants mainly developed calli, whereas bud sprouting occurred in defoliated microcuttings cultured on a modified Murashige and Skoog medium without growth regulators. Isolation and continuous subculture of sprouted buds on the same medium allowed cloning of microcuttings from C. atlantica and C. libani seedlings and bicentennial C. libani trees, thus providing a desirable alternative for multiplying mature trees that have demonstrated superior characteristics. We also report adventitious bud differentiation from isolated embryos of C. atlantica. Neither auxin treatments nor other methods tested, including infection with Agrobacterium rhizogenes, were effective in inducing root initiation.

Cedrus↗

PubMed2002

Genetic transformation of Vitis vinifera via organogenesis.

BACKGROUND: Efficient transformation and regeneration methods are a priority for successful application of genetic engineering to vegetative propagated plants such as grape. The current methods for the production of transgenic grape plants are based on Agrobacterium-mediated transformation followed by regeneration from embryogenic callus. However, grape embryogenic calli are laborious to establish and the phenotype of the regenerated plants can be altered. RESULTS: Transgenic grape plants (V. vinifera, table-grape cultivars Silcora and Thompson Seedless) were produced using a method based on regeneration via organogenesis. In vitro proliferating shoots were cultured in the presence of increasing concentrations of N6-benzyl adenine. The apical dome of the shoot was removed at each transplantation which, after three months, produced meristematic bulk tissue characterized by a strong capacity to differentiate adventitious shoots. Slices prepared from the meristematic bulk were used for Agrobacterium-mediated transformation of grape plants with the gene DefH9-iaaM. After rooting on kanamycin containing media and greenhouse acclimatization, transgenic plants were transferred to the field. At the end of the first year of field cultivation, DefH9-iaaM grape plants were phenotypically homogeneous and did not show any morphological alterations in vegetative growth. The expression of DefH9-iaaM gene was detected in transgenic flower buds of both cultivars. CONCLUSIONS: The phenotypic homogeneity of the regenerated plants highlights the validity of this method for both propagation and genetic transformation of table grape cultivars. Expression of the DefH9-iaaM gene takes place in young flower buds of transgenic plants from both grape cultivars.

Food Industry↗

PubMed2002

Reproducible transformation in two grain legumes--soybean and azuki bean--using different systems.

Two plasmid vectors were introduced into soybean (Glycine max (L.) Merr.) and azuki bean (Vigna angularis Willd. Ohwi and Ohashi) using different transformation systems. Azuki bean epicotyl explants were prepared from etiolated seedlings and co-cultivated with Agrobacterium tumefaciens for 2 days. Adventitious shoots were developed from the callus of the explants on a regeneration medium containing hygromycin, and the shoots were excised and transferred to a rooting medium containing hygromycin at the same concentration. Rooting shoots were transferred to soil and grown in a glass-house to produce viable seeds. PCR analysis confirmed clearly the presence of the hpt gene in most of the azuki beans regenerated under hygromycin selection. A soybean embryogenic suspension culture was generated from immature cotyledons, and used for the introduction of plasmids by particle bombardment. Hygromycin-resistant embryogenic clones were isolated after 8 weeks of hygromycin selection, and then the green clones were matured on the differentiation medium. After desiccation, the embryos were germinated on the rooting medium, and the plants were transferred to soil in a glass-house. More than 50% of the regenerated soybean plants tolerant to hygromycin yielded the hpt fragment on PCR analysis. The azuki bean transformants were obtained more rapidly and with higher efficiency than the soybean transformants.

Fabaceae↗

PubMed1988

Micropropagation of a mature colchicine-polyploid and irradiation-mutant of Betula pendula Roth.

Tetraploid plantlets were regenerated from cultured apical and axillary buds of a 23-year-old colchicine-polyploid and irradiation-mutant Betula pendula Roth tree. Bud explants were grown on modified Murashige and Skoog medium supplemented with 2.0 mg l(-1) benzylaminopurine (BAP) and 0.01 mg l(-1) 1-naphthaleneacetic acid (NAA). The medium allowed both induction of adventitious buds and development of shoots. The cut ends of new shoots produced new buds and shoots during a 4-week culture period. The micropropagated shoots were rooted on modified Murashige and Skoog medium containing 0.1 mg l(-1) NAA as the sole growth regulator. Plantlets were transferred to a peat/soil mixture (1:1) in the greenhouse, acclimated and then transplanted to a cold frame. The regenerated plantlets had a tetraploid chromosome set (4n = 56) and an altered leaf morphology typical of colchicine-polyploid birches. The leaves were hypertrophied and asymmetrical, with curly leaf margins. The mutant nature of the parent tree was also evident in the light-green color of the leaves of the plantlets.

Journal Article↗

PubMed2006

Clonal propagation of softwoods.

Softwoods or gymnosperms, which make up 60% of the forested areas of the world, are economically important as a source of lumber, pulp, and paper. Reforestation is a major activity worldwide and the potential benefits of using clonal planting stock have long been recognized. Tissue culture clonal methods or micropropagation is a newer approach that can be achieved by enhancing axillary bud breaking, production of adventitious buds (organogenesis), and somatic embryogenesis. Plantlet production via organogenesis requires at least four stages: (1) bud induction on the explant, (2) shoot development and multiplication, (3) rooting of developed shoots, and (4) hardening of plantlets. Similarly, the production of plantlets via somatic embryogenesis, which has the potential to produce a larger number of plantlets, and in a shorter period of time, also requires several stages. These include (1) induction, maintenance, and proliferation of embryogenic tissue; (2) maturation (both morphological and physiological) of somatic embryos; and (3) germination and conversion of the somatic embryos. In this chapter, plantlet production via organogenesis from seedling and adolescent/mature explants and somatic embryogenesis from immature and mature seeds of white spruce (Picea glauca) are outlined.

Culture Media↗

PubMed2006

In vitro regeneration of Salix nigra from adventitious shoots.

Black willow (Salix nigra Marsh.) is the largest and only commercially important willow species in North America. It is a candidate for phytoremediation of polluted soils because it is fast-growing and thrives on floodplains throughout eastern USA. Our objective was to develop a protocol for the in vitro regeneration of black willow plants that could serve as target material for gene transformation. Unexpanded inflorescence explants were excised from dormant buds collected from three source trees and cultured on woody plant medium (WPM) supplemented with one of: (1) 0.1 mg l(-1) thidiazuron (TDZ); (2) 0.5 mg l(-1) 6-benzoaminopurine (BAP); or (3) 1 mg l(-1) BAP. All plant growth regulator (PGR) treatments induced direct adventitious bud formation from the genotypes. The percentage of explants producing buds ranged from 20 to 92%, depending on genotype and treatment. Although most of the TDZ-treated inflorescences produced buds, these buds failed to elongate into shoots. Buds on explants treated with BAP elongated into shoots that were easily rooted in vitro and further established in potting mix in high humidity. The PGR treatments significantly affected shoot regeneration frequency (P < 0.01). The highest shoot regeneration frequency (36%) was achieved with Genotype 3 cultured on 0.5 mg l(-1) BAP. Mean number of shoots per explant varied from one to five. The ability of black willow inflorescences to produce adventitious shoots makes them potential targets for Agrobacterium-mediated transformation with heavy-metal-resistant genes for phytoremediation.

2-Aminopurine↗

PubMed2005

Leaf development in the absence of a shoot apical meristem in Zeylanidium subulatum (Podostemaceae).

BACKGROUND AND AIMS: The Podostemaceae are a family of unusual aquatic angiosperms that live in rapids and waterfalls. To adapt to such extreme habitats, the family shows unusual morphologies. This study investigated the developmental anatomy of the shoot of Zeylanidium subulatum borne on the prostrate root attached to submerged rock surfaces. * METHODS: Shoots of Z. subulatum were observed under the microscope using resin-sections. * KEY RESULTS: The shoot has no shoot apical meristem (SAM) and, without it, forms leaves distichously dorsiventrally facing the immediately older leaf. A new leaf forms on the adaxial side of a pre-existing leaf and also on the abaxial side of a leaf on flowering shoots. In both cases, the young leaf is endogenous below the older leaf and maintains histological continuity with it. Shortly after internal initiation, the leaf primordia become separate from each other due to cleavage between adjacent leaves of opposite ranks. The cleavage is caused by intercellular separation as well as by degeneration of vacuolated cells. Loss of the SAM is probably linked with the speculated shift of the site of leaf formation to the root. * CONCLUSIONS: The 'shoot' of Z. subulatum is characterized by the absence of a SAM, endogenous leaf formation in the absence of a SAM, cleavage between leaf primordia, and adventitious leaf formations. These innovations occur in some Podostemaceae that have become increasingly adapted to extreme aquatic habitats.

Magnoliopsida↗

PubMed1987

Pleomorphic lipoma of the dermis.

A case of pleomorphic lipoma is described. The neoplastic tissue developed exclusively within the dermis and characteristically involved the fibrous root sheaths of hair follicles, where several layers of lamellar collagen encircled the follicles and entrapped atypical spindle and multinucleated cells. A possible origin of this tumour in the adventitial dermis is postulated.

Aged↗

PubMed1997

Perivascular sensory nerve Ca2+ receptor and Ca2+-induced relaxation of isolated arteries.

The present study tested two hypotheses: (1) that a receptor for extracellular Ca2+ (Ca2+ receptor [CaR]) is located in the perivascular sensory nerve system and (2) that activation of this receptor by physiological concentrations of extracellular Ca2+ results in the release of vasodilator substance that mediates Ca2+-induced relaxation. Reverse transcription-polymerase chain reaction using primers derived from rat kidney CaR cDNA sequence showed that mRNA encoding a CaR is present in dorsal root ganglia but not the mesenteric resistance artery. Western blot analysis using monoclonal anti-CaR showed that a 140-kD protein that comigrates with the parathyroid CaR is present in both the dorsal root ganglia and intact mesenteric resistance artery. Immunocytochemical analysis of whole mount preparations of mesenteric resistance arteries showed that the anti-CaR-stained perivascular nerves restricted to the adventitial layer. Biophysical analysis of mesenteric resistance arteries showed that cumulatively raising Ca2+ from 1 to 1.25 mol/L and above relaxes precontracted arteries with an ED50 value of 2.47+/-0.17 mmol/L (n=12). The relaxation is endothelium independent and is unaffected by blockade of nitric oxide synthase but is completely antagonized by acute and subacute phenolic destruction of perivascular nerves. A bioassay showed further that superfusion of Ca2+ across the adventitial surface of resistance arteries releases a diffusible vasodilator substance. Pharmacological analysis indicates that the relaxing substance is not a common sensory nerve peptide transmitter but is a phospholipase A2/cytochrome P450-derived hyperpolarizing factor that we have classified as nerve-derived hyperpolarizing factor. These data demonstrate that a CaR is expressed in the perivascular nerve network, show that raising Ca2+ from 1 to 1.25 mol/L and above causes nerve-dependent relaxation of resistance arteries, and suggest that activation of the CaR induces the release of a diffusible hyperpolarizing vasodilator. We propose that this system could serve as a molecular link between whole-animal Ca2+ balance and arterial tone.

Amino Acid Sequence↗

PubMed2004

[Study on tissue culture of Ficus hirta].

The tissue culture of Ficus hirta Vahl. was studied. The nodes were used as explants and 1/2MS media with different plant growth regulators were tested. The result showed that the adventitious bud differentiation medium was 1/2MS + BA1.0 mg/L + NAA1.0 mg/L and 1/2MS + BA1.5 mg/L + NAA0.5 mg/L, the media for multiplication was 1/2MS +6 - BA0.5 mg/L, and the medium for rooting was 1/2MS + IBA1 mg/L. The cause of nigrescence in the tissue culture and its preventive methods were also discussed in this paper.

Ascorbic Acid↗

PubMed2005

The Agrobacterium rhizogenes rolC-gene-induced somatic embryogenesis and shoot organogenesis in Panax ginseng transformed calluses.

Expression of the Agrobacterium rhizogenes rolC gene in Panax ginseng callus cells results in formation of tumors that are capable to form roots. The selection of non-root forming tumor clusters yielded the embryogenic 2c3 callus line, which formed somatic embryos and shoots independently of external growth factors. Although the 2c3 somatic embryos developed through a typical embryogenesis process, they terminated prematurely and repeatedly formed adventitious shoot meristems and embryo-like structures. A part of the shoots and somatic embryos formed enlarged and fasciated meristems. This is the first indication of the rolC gene embryogenic effect and, to our knowledge, the first indication that a single gene of non-plant origin can induce somatic embryogenesis in plants.

Bacterial Proteins↗

PubMed2006

Developmental and hormonal regulation of direct shoot organogenesis and somatic embryogenesis in sugarcane (Saccharum spp. interspecific hybrids) leaf culture.

Rapid and efficient in vitro regeneration methods that minimise somaclonal variation are critical for the genetic transformation and mass propagation of commercial varieties. Using a transverse thin cell layer culture system, we have identified some of the developmental and physiological constraints that limit high-frequency regeneration in sugarcane leaf tissue. Tissue polarity and consequently the orientation of the explant in culture, size and developmental phase of explant, and auxin concentration play a significant role in determining the organogenic potential of leaf tissue in culture. Both adventitious shoot production and somatic embryogenesis occurred on the proximal cut surface of the explant, and a regeneration gradient, decreasing gradually from the basal to the distal end, exists in the leaf roll. Importantly, auxin, when added to the culture medium, reduced this spatial developmental constraint, as well as the effect of genotype on plant regeneration. Transverse sections (1-2 mm thick) obtained from young leaf spindle rolls and orienting explants with its distal end facing the medium (directly in contact with medium) are critical for maximum regeneration. Shoot regeneration was observed as early as 3 weeks on MS medium supplemented with alpha-naphthalenencetic acid (NAA) and 6-benzyladenine, while somatic embryogenesis or both adventitious shoot organogenesis and somatic embryogenesis occurred on medium with NAA and chlorophenoxyacetic acid. Twenty shoots or more could be generated from a single transverse section explant. These shoots regenerated roots and successfully established after transplanted to pots. Large numbers of plantlets can be regenerated directly and rapidly using this system. SmartSett, the registered name for this process and the plants produced, will have significant practical applications for the mass propagation of new cultivars and in genetic modification programs. The SmartSett system has already been used commercially to produce substantial numbers of plants of orange rust-resistant and new cultivars in Australia.

Cell Polarity↗

PubMed2003

[Study on plant tissue culture of Cornus officinalis].

OBJECTIVE: To set up the optimums of tissue culture for Cornus officinalis. METHOD: A section of stems of the fine varieties of Cornus officinalis were strictly sterilized, cultivated and geminated in medium with addition of different kinds of hormone. RESULT AND CONCLUSION: The WPM medium with 6-BA 2.0 mg.L-1 + ZT 0.1 mg.L-1 + NAA 0.1 mg.L-1 was the optimum medium for the multiplication of adventitious buds. The WPM medium with 6-BA 1.0 mg.L-1 + ZT 0.1 mg.L-1 + GA3 0.5 mg.L-1 + NAA 0.1 mg.L-1 was the optimum medium for cultivation in strength. The plantlet could root well in 1/2 MS medium with IBA 1.0 mg.L-1 + 6-BA 0.1 mg.L-1.

Cornus↗

PubMed2002

A pectin glucuronyltransferase gene is essential for intercellular attachment in the plant meristem.

Intercellular attachment is an essential process in the morphogenesis of multicellular organisms. A unique mutant, nolac-H18 (nonorganogenic callus with loosely attached cells), generated by T-DNA transformation using leaf-disk cultures of haploid Nicotiana plumbaginifolia, lost the ability to form tight intercellular attachments and adventitious shoots. The gene tagged with T-DNA, named NpGUT1 (glucuronyltransferase 1), was similar to the gene for the catalytic domains of animal glucuronyltransferases and was expressed predominantly in shoot and root apical meristems. The transformation of NpGUT1 complemented the nolac-H18 mutation, and the expression of antisense NpGUT1 RNA produced crumbled shoots. The mutation caused defects in the glucuronic acid of rhamnogalacturonan II of pectin, which drastically reduced the formation of borate cross-linking of rhamnogalacturonan II. NpGUT1, which encodes a unique glucuronyltransferase, is a glycosyltransferase gene identified in pectin biosynthesis and is essential for intercellular attachment in plant meristems and tissues.

Amino Acid Sequence↗

PubMedDate not supplied

Genetic variability of somatic embryogenesis in tissue cultures of sugar beet breeding lines.

Genetic variability of callus initiation and plant regeneration has been investigated among three sugar beet genotypes. It was found that TDZ has a genotype-independent effect on callus initiation and is responsible for more than a two-fold increase in the friable callus induction rate and more than a three-fold increase in the shoot regeneration rate from this callus. Along with the genotype-independent organogenesis, regeneration from callus occasionally went through the process of somatic embryogenesis in a highly genotype-specific manner. Despite fast and uncontrollable conversion of embryos to normal plants, it was possible to select and maintain repetitive embryogenic culture without loosing regeneration and root formation capabilities. Extensive experimenting with medium composition and culture conditions resulted in an optimal medium for maintenance of repetitive embryos. Comparing with BAP, low concentrations of TDZ provide higher level of adventitious shoot formation and do not induce vitrification of tissues.

Beta vulgaris↗