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Liquid Culture Systems for in vitro Plant Propagation

Anne Kathrine Hvoslef-Eide ; Walter Preil (eds.)

Resumen/Descripción – provisto por la editorial

No disponible.

Palabras clave – provistas por la editorial

Plant Sciences; Developmental Biology; Agriculture

Disponibilidad
Institución detectada Año de publicación Navegá Descargá Solicitá
No detectada 2005 SpringerLink

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Tipo de recurso:

libros

ISBN impreso

978-1-4020-3199-1

ISBN electrónico

978-1-4020-3200-4

Editor responsable

Springer Nature

País de edición

Reino Unido

Fecha de publicación

Información sobre derechos de publicación

© Springer 2005

Tabla de contenidos

Potentials for cost reduction in a new model of commercial micropropagation

V.A. Savangikar; Chitra Savangikar; R.S. Daga; Sunil Pathak

A commercial micropropagation facility, using semi-solid medium in jam bottles as culture containers and a conventional growthroom (with filtration of the air condition system and artificial lighting), was adapted to a new system which uses proprietary liquid medium, improved handling methods to speed up inoculations in polybags and biologically clean stock cultures. Diffused sunlight in the greenhouse is used as light source for the multiplication and rooting of cultures. The temperature of the greenhouse was only controlled by fan-and pad cooling when exceeding 37°C in the afternoons of hot days, allowing ambient temperature of the greenhouse as sufficient control for the cultures. A representative comparison from the daily commercial production records of both the systems was conducted. In the new system, number of propagules produced per workstation per day increased 5 times, and rate of multiplication of propagules per culture in a multiplication cycle increased 3.5 times. Thus, net improvement of performance of the new system is product of these, i.e. 17 times. This is without infusion of automation and with simultaneous reduction in capital costs, running costs as well as interest cost. Further, losses from contamination and during hardening stage is also reduced in the new system. Cost data has been presented on a comparative basis to avoid disclosing commercially sensitive absolute figures. Use of natural light and ambient temperature for culture incubation achieved about a 75% reduction in capital cost as well as in the cost of electricity. Production of cultures per worker, on account of new proprietary handling methods, was 70% more in the new model, a factor relevant to developed economies as this feature can be utilized in their present systems too, whether rest of the features of new system are adapted or not. Potential reduction, extrapolated from actually achieved results, in cost of production is 92 – 98% in Indian context and is estimated to offer about 55–87% for developed countries, depending upon efficiency of hardening. Thus this model is relevant to developing as well as developed countries. Further scope exists in improvement in cost reduction in tropical and semi-tropical climate. In temperate climate, scope is lesser for cost reduction, but the same may yet be practically worthwhile when compared to costs in conventional model. Over and above the advantages mentioned above, sturdiness and survival of plants in hardening was better in plants coming from new system.

IV. - Commercial Process Development and Culture Environment | Pp. 403-414

A new approach for automation: Sorting and sowing dehydrated somatic embryos of and using seed technologies

J.P. Ducos; B. Florin; J.M. Dupuis; V. Pétiard

Somatic embryos of L. and L. (var. Robusta) were dehydrated under a 43 % relative humidity then placed in the hopper of a precision seeding system used in the transplant industry. The seeder was adjusted to distribute the embryos onto horticultural trays, each one containing 240 cells filled with soil. As a preliminary result, 72 % and 88 % of the individual cells received a single embryo, in Daucus and Coffea respectively. The embryo-to-plantlet conversion rate was not affected either by the vibration of the hopper or by the nozzles. In carrot 66 % of the embryos germinated after the use of the seeding system (62% germination for the control). Sorting methods traditionally used for the seeds (e.g. air column, vibrating table) can also be used. Such an approach, based on desiccation as a key step, has the potential for a complete automation of the large-scale handling and delivery of somatic embryos.

IV. - Commercial Process Development and Culture Environment | Pp. 415-423

Use of the temporary immersion bioreactor system (RITA®) for production of commercial clones in Mondi Forests (SA)

B. Mc Alister; J. Finnie; M.P. Watt; F. Blakeway

In order to optimise tissue culture systems and to meet production targets, Mondi Forests’ biotechnology programme has in the last two years concentrated efforts on the use of the RITA® temporary immersion bioreactor system. Protocols have been established for six clones. Results indicate a four- to six-fold increase in yield, in half the time, with the RITA® system when compared with axillary bud proliferation on semi-solid media. Furthermore, plants produced from the RITA® system are hardier and acclimatize better, giving higher yields of hardened-off plants. The establishment of aseptic axillary shoots into the RITA® system is from shoots in the semi-solid system. Highest multiplication was achieved using 30-second flushes of medium every 10 minutes, starting with 50 shoots per vessel. The multiplication cycles in RITA® are between 14 and 18 days, compared with 25 to 28 days in a semi-solid system. There is minimal callus evident on the leaves and bases of the stems of plants in the RITA® system and, in addition, cold-tolerant plants have a greater rooting competence when compared with plants coming from the semi-solid system. rooting of RITA®-derived plantlets is substantially better than the plants from the semi-solid media.

IV. - Commercial Process Development and Culture Environment | Pp. 425-442

Efficiency in thin-film liquid system for micropropagation

Jeffrey Adelberg

Three varieties of (‘striptease’, ‘Minuteman’ and ‘stiletto’) at four densities (40, 80, 120 and 200 explants per litre) were micropropagated on semi-solid agar and a thin-film liquid system with intermittent wetting of plant tissue. The mechanics of wetting by a small wave front required a larger rectangular vessel (11 × 27 = 297 cm) compared to the common cylindrical baby food jar (18 cm). Plants multiplied more rapidly in the agitated thin-film system than on agar. Lower plant densities increased rates of multiplication in liquid, but had little or no effect on multiplication rate on agar. Increasing plant density lowered the overall multiplication rate, but yielded greater numbers of plants per vessel. Yield, tabulated for utilization of shelf-space in growth room, was greater at all densities in rectangular vessels of liquid than conventional jars of agar media. Increased plant density lowered the sugar residual in media following the culture cycle and liquid media had less residual sugar than agar media. A liquid medium with 50g l sucrose was concentrated enough so that sugar depletion did not limit growth, even at the highest densities. The liquid system allows the technician to skip the step of manually spacing and orienting the freshly cut bud tissue at the transfer station. Harvesting 75 – 100 plants per vessel from the large rectangular vessels resulted in most efficient use of technician time. Plants from liquid and agar acclimatized to greenhouse. Increased multiplication, space utilization, sugar availability and worker efficiency was demonstrated to be greater in thin-film liquid than more conventional agar-based system.

IV. - Commercial Process Development and Culture Environment | Pp. 443-457

Aeration stress in plant tissue cultures

Michael B. Jackson

The requirement for both sterility and the avoidance of dehydration in plant tissue cultures can impose sealing requirements that severely limit the rate of gas exchange in and out of the culture vessel. Conditions within the culture vessel, such as the depth of any water cover, the presence of gelling agents, the bulk and porosity of the tissue and the temperature, also strongly influence rates of gas exchange, primarily driven by diffusion. This article uses elements of Fick’s Law of Diffusion to identify key factors limiting gas exchange between a culture and its immediate surroundings. In particular, it identifies static liquid media, gelling agents, large tissue mass and warm temperatures as imposing severe limits on diffusive flux for gases such as O, CO and ethylene. The principal barrier to diffusive exchange of gases between the and atmospheres is the wall of the enclosing vessel. This is invariably made of glass or plastic that is gas-impermeable and well-sealed against evaporative drying or entry of micro-organisms. Cultures enclosed in this way will, inevitably, asphyxiate unless a compensating pathway for diffusive gas exchange is contrived or replaced by some system of convective flow that carries gases to and from the tissue. Supplementing diffusive aeration with convective flow is the basis of most successful hydroponics systems for whole plants and may be a prerequisite for securing levels of aeration suitable for autotrophic cultures. The paramount consideration is the extent towhich the total rate of consumption or production of a particular gas by the cultured tissues is matched by the maximum rates of gas transport imposed by the culture itself, its immediate surroundings and the ventilation and sealing system of the culture enclosure.

IV. - Commercial Process Development and Culture Environment | Pp. 459-473

Macro- and micronutrient nutrition of plants in greenhouses, hydroponic systems, and culture on gelled media

Hans R. Gislerød; Ravichandran Selliah; Kwadwo Owusu Ayeh; Anne Kathrine Hvoslef-Eide

Nutrition for and greenhouse production systems is reviewed and found to be broadly similar. The optimal pH (5.0–6.0) is independent of the growing system used, with some lower or higher pH values required for special crops. The pH of nutrient solutions is regulated either with acid or via the NH:NO ratio. For solutions, it is possible to use buffers (MES or TRIS), while in greenhouse systems the pH buffering capacity relies mostly on the quantity of colloids in the growing medium.

The normal nutrient solution conductivity for vegetative propagation is 0.8–1.2 mS cm and 1.5–2.0 mS cm for growth in greenhouse production. In micropropagation on gelled media the conductivity will be between 3.0–6.5 mS cm. When spraying nutrient directly on the leaves in greenhouse production either to increase uptake of a particular element or to prevent a deficiency, the same concentration as for micropropagation should be used.

The air humidity will usually be greater in micropropagation systems than in greenhouse production. In culture of plant parts, uptake of the different nutrient elements will be mainly by diffusion. When comparing the proportion between and growing systems, there is a surprisingly low content of Ca and P for micropropagation. For micronutrients, micropragation media are low in Fe and Cu content and high in Mn and Zn content, compared to that required for greenhouse production. Light has been found to have an effect on the stability of iron-chelates and thus on the quantity of iron available in the media for uptake. We present results to show the importance of the right nutrient content of the medium and how it affects growth and development , but mostly the increase in development rate to flowering.

IV. - Commercial Process Development and Culture Environment | Pp. 475-492

Adaptions of the mineral composition of tissue culture media on the basis of plant elemental analysis and composition of hydroponic substrates

Han Bouman; Annemiek Tiekstra

For the improvement of propagation protocols of and , we adapted the macronutrients according to the elemental composition of adult leaves. In comparison with normally used media, in particular, the Ca and P concentrations were much higher. Using the adapted media, for both crops growth was much improved. The relative concentrations of macronutrients in these modified media formulations is much more in accordance with the mineral composition of nutrient solutions used in hydroponic cultures. For , we also made adaptations to the micronutrients according to the elemental composition of nutrient media in hydroponics (, increased Cu-content and decreased Mn-content). This resulted in additional improvement of growth; by this adaptation, the plants became larger and also greener (only with the Cu adaptation).

IV. - Commercial Process Development and Culture Environment | Pp. 493-505

Development and validation of an efficient low cost bioreactor for furanocoumarin production with shoot cultures

E. Gontier; S. Piutti; A. Gravot; S. Milesi; A. Grabner; B. Massot; K. Lievre; M. Tran; J.L. Goergen; F. Bourgaud

Despite efforts made to produce plant secondary metabolites from cell suspensions, only a few industrial applications have been successful. Generally, higher yields are obtained when cultivating organs (roots or leafy stems) instead of undifferentiated cells. In this case, new problems arise because of the structure of the plant material, and special bioreactors have to be built for such applications. Furthermore, the high cost of commercial bioreactors may limit the number available for the researcher to carry out many experiments in parallel. Because of this, we developed a very low cost system (i.e; bioreactors) that allows good growth of L. shoots and production of secondary metabolites (. furanocoumarins). The development of a very simple auto-priming siphon allows the use of common jars ranging from 3 to 20 litres for temporary immersion cultures. The very low cost of such a home-made bioreactor allows scientists to run many different experiments at the same time. It thus saves time in optimising the culture medium parameters and in replicating trials before reaching the step of final culture system development with highly equipped (costly) bioreactors.

V. - Biomass for Secondary Metabolite Production | Pp. 509-524

Comparison of secondary plant metabolite production in cell suspension, callus culture and temporary immersion system

Dirk Wilken; Elio Jiménez González; Annette Hohe; Miguel Jordan; Rafael Gomez Kosky; Guillermo Schmeda Hirschmann; André Gerth

Cell and organ cultures of and were established for the production of secondary metabolites . Shoot multiplication was performed by conventional micropropagation on agar-solidified medium as well as in temporary immersion systems (TIS), the latter resulted in higher multiplication rates compared to the culture in microcontainers for all plant species tested. The concentration of bioactive compounds was determined in different cell and organ cultures and was compared to field grown plants. For the highest content of rosmarinic acid was found in cell cultures, for the other three species in field grown plants. Concentrations of bioactive compounds were always higher in plant material grown in TIS compared to cell suspension and callus cultures.

V. - Biomass for Secondary Metabolite Production | Pp. 525-537

Cultivation of root cultures of in different bioreactors and in temporary immersion — Comparison of growth and saponin production

Tomáš Vaněk; Lenka Langhansová; Petr MaršÍk

Different systems of large-scale cultivation of multiple adventitious roots of Panax ginseng C. A. Meyer were compared to cultivation in Erlenmeyer flasks. Adventitious roots were isolated from plantlets regenerated from somatic embryos and cultivated separately in liquid media. Multiplication of adventitious roots was performed in liquid Schenk and Hildebrandt (1972) medium containing 3% sucrose, and 24.6 µmol indole-3-butyric acid. The highest saponin content of 28.51 mg g of dry weight was found in adventitious roots cultivated in the RITA temporary immersion system (TIS). The best production of biomass was achieved in RITA vessels and standard Erlenmeyer flasks placed on rotary shaker, followed by the Applikon 3-litre bioreactor and a simple airlift reactor. Saponin production in Erlenmeyer flasks was 10.07 mg g of the dry weight while the production in the Applikon 3-litre bioreactor was only 3.60 mg g. Other bioreactor systems tested showed neither significant saponin production nor high biomass production.

V. - Biomass for Secondary Metabolite Production | Pp. 539-546