Temporary immersion systems in plant micropropagation

Centre de Coopération Internationale en Recherche Agronomique pour le Développement – Amis (CIRAD-AMIS), CIRAD
Plant Cell Tissue and Organ Culture (Impact Factor: 2.61). 05/2002; 69(3):215-231. DOI: 10.1023/A:1015668610465

ABSTRACT Temporary immersion systems for plant micropropagation have been described and grouped into 4 categories according to operation: tilting and rocker machines; complete immersion of plant material and renewal of the nutrient medium; partial immersion and a liquid nutrient renewal mechanism; complete immersion by pneumatic driven transfer of liquid medium and without nutrient medium renewal. The positive effects of temporary immersion on micropropagation are indicated for shoot proliferation and microcuttings, microtuberization and somatic embryogenesis. Immersion time, i.e. duration or frequency, is the most decisive parameter for system efficiency. Optimizing the volume of nutrient medium and the volume of the culture container also substantially improves efficacy, especially for shoot proliferation. Temporary immersion also generally improves plant material quality. It results in increased shoot vigour and in the frequency of morphologically normal somatic embryos. Hyperhydricity, which seriously affects cultures in liquid medium, can be eliminated with these culture systems or controlled by adjusting the immersion times. Plant material propagated by temporary immersion can perform better during the acclimatization phase than material obtained on semi-solid or in liquid media. Successful regeneration of plants, after direct sowing on soil of Solanum tuberosum microtubers and Coffea arabica somatic embryos produced in temporary immersion bioreactors, has been demonstrated. As could be expected when using liquid medium for micropropagation, several estimations confirm large gains in efficacy from temporary immersion. The parameters most involved in reducing production costs include: (1) the drastic reduction in work; (2) reduction in shelving area; (3) reduction in the number of containers used; (4) better biological yields. Scaling-up somatic embryogenesis and shoot proliferation procedures involving temporary immersion systems in order to commercialize this process are now taking place.

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    ABSTRACT: Castilleja tenuiflora, a species highly valued for its medicinal properties, is threatened in the wild. We evaluated the effects of six different immersion cycles in a temporary immersion bioreactor on C. tenuiflora shoot growth, proliferation rate, phenolics content, flavonoid content, and antioxidant activity. We also evaluated the regeneration capacity of the shoots. The highest proliferation rate (nine shoots per explant) was obtained using an immersion cycle of 5 min every 12 h, and the longest shoots (38.8 +/- 1.9 mm) were obtained using an immersion cycle of 5 min every 24 h. Shoots obtained from immersion cycles of 30 min every 24 h or 5 min every 24 h showed 100% rooting efficiency. Shoots obtained from immersion cycles of 30 min every 3 h or 30 min every 12 h accumulated H2O2, developed abnormal stomata, and showed symptoms of hyperhydricity. These characteristics were associated with a low survival rate (16-80%) when the plants were transferred to potting mix. The shoots from an immersion cycle of 30 min every 24 h showed the highest total phenolics content, which coincided with the highest antioxidant activity in the 2,2'-azinobis (3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS) free-radical scavenging assay (161.74 +/- 10.06 mu mol Trolox/g dry weight (DW)). The shoots from an immersion cycle of 5 min every 24 h showed the highest activity in the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free-radical scavenging assay, and those from an immersion cycle of 5 min every 3 h showed the strongest reducing power. These results show that temporary immersion culture represents a reliable and efficient method for in vitro micropropagation of C. tenuiflora.
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    ABSTRACT: Plant density was varied with P, Ca, Mg, and KNO3 in a multifactor experiment to improve Curcuma longa L. micropropagation, biomass and microrhizome development in fed-batch liquid culture. The experiment had two paired D-optimal designs, testing sucrose fed-batch and nutrient sucrose fed-batch techniques. When sucrose became depleted, volume was restored to 5% m/v sucrose in 200 ml of modified liquid MS medium by adding sucrose solutions. Similarly, nutrient sucrose fed-batch was restored to set points with double concentration of treatments' macronutrient and MS micronutrient solutions, along with sucrose solutions. Changes in the amounts of water and sucrose supplementations were driven by the interaction of P and KNO3 concentrations. Increasing P from 1.25 to 6.25 mM increased both multiplication and biomass. The multiplication ratio was greatest in the nutrient sucrose fed-batch technique with the highest level of P, 6 buds/vessel, and the lowest level of Ca and KNO3. The highest density (18 buds/vessel) produced the highest fresh biomass at the highest concentrations of KNO3 and P with nutrient sucrose fed-batch, and moderate Ca and Mg concentrations. However, maximal rhizome dry biomass required highest P, sucrose fed-batch, and a moderate plant density. Different media formulations and fed-batch techniques were identified to maximize the propagation and storage organ responses. A single experimental design was used to optimize these dual purposes.
    PLoS ONE 01/2015; 10(4):e0118912. DOI:10.1371/journal.pone.0118912 · 3.53 Impact Factor
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    ABSTRACT: Currently in vitro plantlets and microtubers provide the basis for pre-base production of potato seeds, from which minitubers are produced under covers - they serve later as seed material to be planted in the field. The aim of the research was to determine the possibility for multiplication of material produced in vitro directly in field conditions. The research assessed PVY, PVM and PLRV infection of potato tubers derived from plants grown directly from in vitro plantlets, microtubers, minitubers and traditional seed potatoes planted in the field at different times. Moreover, testing in laboratory conditions, the susceptibility of these plants to virus infection was determined for the case of artificial inoculation of Myzus persicae and Aphis nasturtii. It was found that the infection of tubers derived from in vitro plantlets and microtubers was greater than that of seed potatoes and minitubers. Yet it seems that the reason for their higher infection level resulted not from the plant's sensitivity or its greater attractiveness to aphids but from a largely unknown cause. Earlier planting of microtubers and in vitro plantlets in the field in case of the more resistant cultivar and certainly later in relation to the main time of planting had an impact on limiting the PVY and PVM infection of potato tubers. Hence multiplication of microtubers and in vitro plantlets in field conditions could be very economical using cultivars which are relatively resistant to viruses. However, adopting a later than usual planting period (end of June) and applying an additional protective cover (such as non-woven agricultural fabric) in the first period of a plant's growth, promotes multiplication of microtubers and in vitro plantlets in field conditions for cultivars with low resistance levels.
    American Journal of Potato Research 10/2014; 91(5):554-565. DOI:10.1007/s12230-014-9388-6 · 0.95 Impact Factor


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May 15, 2014