Potato is one of the most important crops worldwide. Genetic resources of potato (Solanum tuberosum L. ssp. tuberosum) and related cultivated species are conserved through storage of tubers, in vitro plants and in cryopreservation. Cryopreservation, storage in or above liquid nitrogen, is the best option to maintain vegetatively propagated plants in the long term. The present review gives comprehensive information about various cryopreservation techniques for potato published from 1977 until the present. It discusses factors that affect the process and success of cryopreservation, such as donor culture conditions, preculture, cooling, warming and post-culture treatments. Studies are presented that analyse the histological and ultrastructural changes after different cryopreservation steps and the morphological pathways during regeneration of plants after rewarming. The maintenance of genetic stability in potato after cryopreservation has also been demonstrated by various phenotypic and molecular methods. The first thermal analyses on potato shoot tips are presented using differential scanning calorimetry to analyse the state of water during cooling and warming. Biochemical analyses of different compounds, such as soluble sugars and proteins, have been performed to understand and improve existing cryogenic methods. Potato is an example where successful virus elimination has been obtained via cryopreservation of shoot tips (cryotherapy). There are already cryopreserved collections of potato shoot tips in

Experiments were initiated to develop protocols for four species of Dioscorea, namely D. bulbifera L., D. oppositifolia L., D. alata L. and D. cayenensis Lam., and to assess the effects of various factors on the survival and regrowth of explants after rewarming. The treatments consisted of three different methods-vitrification, droplet and a modified droplet method-as well as the following variables-cryoprotective solution, sucrose concentration in the preadaptation medium and cold acclimation. While most of the protocols resulted in low to zero survival and regrowth rates, maximum survival rates of 100% and 75% were obtained for D. oppositifolia L. and D. cayenensis Lam., respectively, using two different protocols of the modified droplet method. A higher average survival rate was obtained using the droplet and modified droplet techniques than for the original vitrification methods. When the average of all the experiments was taken, more than one-half (52%) of the surviving explants developed further within 1 month with the modified droplet method compared to zero regrowth with the two other methods. The optimum protocol for cryopreservation is specific for each genotype.

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Two experiments were conducted with four accessions of mint (Mentha spp.) on MS medium for their in vitro performance. In the first experiment apical and nodal explants were cultured at both 20°C and 25°C. Data was recorded at second and at fifth week. Both apical and nodal explants of mint showed better leaf production at 25°C than 20°C. Nodal explants of mint cultured at 25°C in both cultivation periods exhibited the highest number of leaves. In the second experiment apical explants were cultured in four different culture vessels viz., industrial glass jar (IG), magenta vessel (MV), Erlenmeyer flask (EF) and culture tube with 1(CT1) and 2(CT2) explants at 25°C for 6 weeks. The highest weight loss from the media, evapo-transpiration and fresh weight gain were recorded in IG and next in MV. The lowest weight loss from the media and fresh weight gain both were found in CT2. However the lowest evapo-transpiration was noted in EF. The highest numbers of leaves were recorded from MV. Without explants, depletion of medium and increase of headspace were both higher in IG than in the other vessels. Overall, Magenta vessel GA 7 showed the best in vitro performance.

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.Keywords Solanum tuberosum Á Solanum acaule Á Solanum demissum Á DMSO droplet method Á Carbohydrates Á Amino acids Abbreviations CT Constant temperature preculture (22°C constant temperature 7 days) AT Alternating temperature preculture (22/8°C day/night temperature 7 days)