Influence of alternating temperature preculture on cryopreservation results for potato shoot tips

Cryopreservation is the most suitable long-term storage method for genetic resources of vegetatively maintained crops like potato. In the Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) the DMSO droplet method is applied, and so far more than 1000 accessions are cryopreserved with an average regeneration rate of 58%. New experiments with four potato accessions using alternating temperatures (22/8°C day/night temperature, 8 h photoperiod, 7 d) prior to cryopreservation showed improved regeneration. The influence of this preculture on the shoot tips was studied for two wild, frost resistant species Solanum acaule and S. demissum and for two cultivated, frost sensitive potatoes S. tuberosum ‘Désirée’ and ‘King Edward’. Comparison of liquid and solid media after cryopreservation showed improved regeneration on solid media with higher regeneration percentages, less callus formation and better plantlet structure. In comparative analyses biochemical factors like soluble sugars, starch, and amino acid concentrations were measured. Shoot tips after constant and after alternating temperature preculture were analyzed. Total concentrations of soluble sugars (glucose, fructose, and sucrose) were higher for all accessions after the alternating temperature preculture, which could be the reason for improved cryopreservation results

In vitro storage and cryopreservation as substantial complements in concerted actions to better maintain and use crop germplasm

The existence of modern cultivated plants is a result of interplaying modification by breeding and maintenance of valuable characters by storing the plant biodiversity. Genebanks were established to maintain and provide the crop diversity for use on the long term. Therefore, all possible methods for maintaining plant germplasm are necessary in order to fulfil this purpose for the various types of germplasm (field culture, seed storage, in vitro culture and cryopreservation). In vitro culture and cryopreservation are the tools with which laboratories can contribute to conservation but also to the improvement of germplasm. They are useful to protect germplasm from threats imposed by environmental hazards, pests and diseases as well as human damage. Some case studies are presented. In case of potato, a crop of high importance, 2700 accessions are maintained in vitro and about 1200 accessions are stored in liquid nitrogen through cryopreservation at the IPK Gatersleben. Also in case of banana, 1250 and 850 accessions are respectively stored in vitro and under cryopreserved conditions at the Bioversity Musa collection, Leuven, Belgium. The interrelationships between field culture, in vitro storage, and cryopreservation are determined by safety and cost factors. Most divers are the collections of Allium and of medicinal plants belonging to the family Lamiaceae, where variable sets of maintenance methods are followed in dependence on the propagation system of the material. Whereas in vitro storage and cryopreservation initially were mainly considered as tools for preservation of clonal crops, they may also contribute to outbreeding populations when they are used complementary to other methods. With the increasing use of these methods on many diverse locations, development of international collaboration becomes a focal point. This involves technology transfer between different partners, joint development of new methods and creation of networks and benefit-sharing larger storage entities. This covers also safety aspects like safety duplication of samples. The COST action 871 "Cryopreservation of crop species in Europe", having joined 21 countries, the European GenRes Project EURALLIVEG, having created a cryobank system of three countries, and the TRUST project on cryopreservation of tropical vegetatively propagated species are discussed as examples. They follow some earlier initiatives such as an EU GenRes project on Allium preservation and the European cryopreservation project CRYMCEPT. Other actions are integrative on a national level such as the CRYOVEG project, recently implemented in France. This project aims at integrating in a rational and coordinated manner cryopreservation in the overall strategies employed for conserving a range of crop species of national interest. In vitro storage and cryopreservation are not simply alternatives designed to replace other methods of conservation but rather valuable complements in an allinclusive strategy to maintain and use plant germplasm in the long term.status: publishe

Allium genetic resources

An overview of the developments in Allium genetic resources during the past 25 years is presented in this chapter. A first important development has been the introduction and further development of web-based genebanking information systems (e.g. GENESYS, PLANTSEARCH), which facilitated the exchange of data to a large extent between Allium collection holders worldwide. These information systems made it possible to obtain an overview of the Allium genetic resources managed worldwide and identify the gaps in collections which still need to be filled, especially in the face of the ongoing genetic erosion. A second important area of progress has been the development of new methods for the maintenance of Allium germplasm, especially cryopreservation. This method has made it possible to maintain Allium accessions in a cheap and effective way. The method is especially important for the conservation of vegetatively maintained germplasm. Other developments in Allium genebanking are the improvement of the health status of the germplasm kept in the collections and the continuing characterization and evaluation of germplasm, which stimulates the utilization of the Allium genetic resources held in genebanks. Significant changes could also be observed with respect to acquisition and exchange of plant genetic resources due to many and complex new regulations on the legal and organizational levels due to the adoption of the CBD and IT-PGRFA by many countries. It makes the handling of the plant accessions safer and more consistent but also more circumstantial. Finally, we need to underline that in an increasingly changing world with all the threats of genetic erosion and extinction due to disappearance of traditional cultivation methods, devastation of our environment and climatic change, the conservation of genetic resources is of prime importance for agriculture. Especially for breeders, a highly diverse genepool of a crop plant is an invaluable treasure. The importance to keep this treasure will no doubt become even more important in the future

Cryobiotechnological Studies in Vanilla: The Orchid of Multi-industrial Uses

In this work we provide new information about several cryobiotechnological studies conducted using Vanilla planifolia species as a case study. We evaluated the effect of three vitrification-based techniques, droplet-vitrification (D-V), V cryo-plate (V-Cp) and D cryo-plate (D-Cp), on the survival of two types of in vitro explants (shoot-tips and root-tips). Using differential scanning calorimetry (DSC), there were defined thermal characteristics of shoot-tips subjected to several desiccation times with SG following D-Cp procedure. It was calculated the removal of osmotically active water (OAW) by the effect of desiccation duration after the osmoprotective steps of preconditioning and loading. Using ISSR markers with six selected ISSR primers, we also evaluated the impact of tissue culture and of cryopreservation on genetic stability of vanilla shoot-tips dehydrated with the vitrification solution PVS2 and comparing D-V and V-Cp procedures. All these investigations are allowing the development and optimization of reliable cryopreservation protocols for long-term storage of Vanilla germplasm.Fil: González Arnao, María Teresa. Universidad Veracruzana; MéxicoFil: Hernández Ramírez, Fabiola. Universidad Veracruzana; MéxicoFil: Dolce, Natalia Raquel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Botánica del Nordeste. Universidad Nacional del Nordeste. Facultad de Ciencias Agrarias. Instituto de Botánica del Nordeste; ArgentinaFil: Rascón Díaz, Martha P.. Universidad Veracruzana; MéxicoFil: Cruz Cruz, Carlos A.. Universidad Veracruzana; Méxic