AN EFFECTIVE PROCESS FOR THE IN VITRO-IN VIVO PRODUCTION OF POTATO MINITUBERS

Abstract: Effective process for the in vitro-in vivo production of potato minitubers in such a way, that in vitro tuber initiation is induced on in vitro rooted plants, and these plants are planted into glasshouses, the growth of haulm is limited by treatment with antigibberellic chemicals, minitubers are collected and plants are transplanted, properly treated and minitubers produced during the vegetation period are collected periodically.

Description: S 1 An effective process for the in vitro-in vivo production of :potatα minitubers Technical field The present invention celates to an effective process for 5 the in vitro-in vivo production of potato minitubers. Background art Before the date of application of this invention our Hungarian patent application no. 1160/84. - not published yet - claims a process for the mass production of potato's 10 propagation material free of viruses and viroids, by plant tissue culture, said process comprises, that the potato's shoottip-tissue cells are cultivated and propagated in vitro on nutritive media, then shoots are rooted Or formation of minitubers is induced, the in vitro rooted plants are plan- 15 ted into glasshouses to grow in- vivo mother plants or in vitr tuber formation can be induced- from these minitubers further mother plants can be grown and from these seedlings or small tubers can be obtained as propagation materials, or the mini¬ tubers can be used as propagation materials. The formation of 20 seedlings or tubers can be directed by influencing the patatine-synthesis of the mother plants. In the mentioned application we discussed in detail the prior art for the production of in vitro propagation material of potato, and it has been found that several valuable partial 25 results are available, no complex system was known which coul be succesfully used for producing an appropriate propagation material on large scale. The mass propagation of potato's propagation material is a little bit difficult, - because of the nature of this plant -. The multiplication rate of potato propagated by t adicio- πal process is low (maximum 10-12 times yearly), so the traditional c ultivar maintenance needs 8-12 years. The presently available cultivars of potato a re susceptible to different fungus-, bacteria- and virus diseases. The tra- ditional cultivar maintenance can be realised only in those countries, where the level of infection is low enough because of favourable oecological factors. The water content of potato's propagational material is approx. 90%, so transportation and storage of it needs special conditions. In a given case the transportation costs can be higher than the value of potato at the place of production. The process disclosed in our Hungarian patent application no. 1160/84 is suitable for the elimination of the above mentione difficulties. Using that process the rate of propagation is hi the time needed for cultivar maintenance can be reduced to 1-2 years, healthy propagation material can be produced, the costs of transportation and storage can be significantly reduced by propagation of minitubers. According to this earlier process potato propagation materials have to be produced in laboratory and glasshouse. Because the costs of investment and maintenance are high - in both cases - so the profitability of the process is dependent on the amount of propagation material produced on a unit area. The foundation of glasshouses is more expensive than the estab¬ lishment of laboratory. The transplantation of plants produced in laboratory needs good technical level in glasshouses (exact soil heating facilities, regulation of humidity, additional illumination Keeping in mind, that plants transplanted from laboratory into glasshouses have to be grown economically and this can be guaranteed in the case of minituber production only by the best utilisation of glasshouses. According to our earlier process 400-800 pcs of minitubers can be produced per squaremeters. This fact limits the distribution of the utilisation of glasshouses for minitube production. We have worked out a new method to increase significantly the efficiency of minituber production per unit area. According to this process of this invention - what is not obvious from the known prior art - the efficiency of the earlier process can be increased at least by 300-500%. Disclosure of the invention The subject of our invention is that disease-free and propagated plants produced by the known tissue culture meth are rooted in liquid media in vitro under sterile laborat circumstances, when the plants have developed roots the nutritive media is changed to a media containing anti¬ gibberellic chemical for the induction of tubers and plants are incubated in this media. After induction of tubers plant are treated with a chemical which promotes the rooting and are planted in solid media into glasshouses. The vegetative growth of plants is limited by decreasing the level of gibberellin to the level of cytokinin content periodically and regularly, and the produced minitubers are collected from roots, than plants treated witn a rooting hormon are planted back into solid media and are treated regularly and periodically with antigibberellic chemical, than the produced minitubers are _ collected again. The process - according to the invention - can be realised that disease-free plants are produced from tissue culture by known methods. The virus- and disease-free plants can be propagated by any of the known tissue culture methods. Plant propagated by tissue culture are rooted in liquid media. After the rooting phase - plants have adjacent roots - the rooting media is changed to a media for tuber induction, which is mainly MS media (Murashige and Skoog: Physiol. Plant. 15. 473-397 /1962/ ) and there is a difference in the amount of inorganic nitrogen source (here it is 10% of the original) and nitrogen requirement of plants is guaranteed by adding organic nitrogen source (glutamine, asparagine) during the induction. The concentration of saccharose in the induction media is higher than in the MS media, 40 g/1 saccharose can be preferably used. The media used for induction contains cytokinins at a high concentration, in average 5-10 mg/1 is suitable. The essentia parts of media used for tuber induction are the different antigibberellins. Such chemicals are coumarin and its deri- vates. chlor-choline-chloride, (CCC), preferably dichloro- -acetyl-hexamethylene-imiπe. This last chemical has growth- -li itiπg and antidoting effects. In plants it has an effect to tuber induction by decreasing the level of gibberellin compared to cytokinins and has an advantageous influence to the structure of cell-membranes of plants. According to the process of this invention dichloro-acetyl-hexamethylene -i ine can be used at a concentration of 6-20 mg/1. Plants are incubated in the induction media under 18-20°C 8 hours illumination for 10-14 days. After the induction potato tubers are developed in solid media in glasshouses. Before transplantation into glass¬ houses induction media is washed out from roots and to promote the rooting and to prevent plants from infection of Rhizoctonia plants are plunged into a liquid containing 0,05 mg/1 iπdole-acetic acid and 0,2% Previcur-N (=propil- -N-/3-di ethyl-amino-propil/-carbamat hydrochloride). After getting rooted the plants in glasshouses - 3 weeks after transplantation - plants are sprayed with a solution containing 10 mg/1 coumarine and/or -5-10 mg/1 dichlor-acety -hexamethylene-imine once a week. So plants remain small bu their physiological stage is suitable for tuber formation. If it is necessary, plants can be pinched back to prevent further growth. After transplantation in glasshouses (1,5-2 months later) the banch of plants is removed from the soil and tubers bigger than 0,5 cm are collected from the roots. Approx. 10 tubers per squaremeter can be collected from plants grown in this manner at the first time. Roots can be damaged during the removal of plants and tuber The banch of plants is plunged again into a solution contai ing 0,05 mg/1 indole-acetic acid and 0,2% Previcur-N, than planted into the original or new soil. After the rooting phase the earlier treatments are repeated (10 mg/1 coumariπ and/or 6 mg/1 dichlor-acetyl-hexamethylene-imine) . The re¬ moval of banches of plants and tubers, treatment with the above written solution, transplantation of plants can be repeated after 3-4 weeks. The second harves of tubers can be 2-3000 pcs/squaremeter. When plants are ageing - 4 months after planting - those are removed and destroyed. Plants treated in the above written manner can produce 2500-4000 pcs tubers/squaremeter dependently on cultivars, and this amount is 3-4 times more than one can get by the earlier method. The collected tubers are suberised for two weeks, under diffuse light and 70% humidity, than are grading and stored at 2-5°C. This process according to the invention can be repeated 2,5 times yearly, it means that the efficiency of minituber -production is increased 4-5 times of the earlier one. Best mode of carrying out the invention The following example illustrates the invention: Example The "Somogy Gyongye" potato cultivar is propagated by tissue culture on a known way. After getting the proper plant numbe shoots are rooted in liquid media. After two weeks rooting the liquid media is changed to another one for tuber inducti The composition of media for tuber induction: Murashige and Skoog (MS) basic media, 1/10 concentration, NH.N0-, and KNO- j , 20 mg/1 glutamic acid, 40 g/1 saccharose, 5 mg/1 benzyladeni 25 mg/1 coumariπe and 6 mg/1 dichloro-acetyl-hexamthylene- -imine. Plants are incubated in this media for tuber inducti for 10 days, under 8 hours 200 lux/m illumination, at 18°C. After the induction bunches of plants are remove from culture-flasks, plants are washed in tap water and snaked for the removal of excess water and plunged into a solution containing 0,08 mg/1 indole-acetic acid and 0,2% Previcur-N, than planted into peat mixture, 50 bunches of plants/m . After rooting plants small ridge formation is done (2 cm), than from the second weeks after planting pla are sprayed with a solution containing 10 mg/1 coumarine a 6 mg/1 dichloro-acetyl-hexamethylene-imine once a week. Af planting - 1,5-2 months later - bushy bunches of plants ar removed from the soil on growing-beds. Tubers bigger than 0,5 cm are collected with a comb made for this purpose or by hand. Peat and soil are shaked from the removed bunches plants, which are plunged into a solution containing 0,08 indole acetic acid and 0,2% Previcur-N, than are planted back to the original soil and are irrigated. The treatment of plants is continued by spraying them with a solution co taining 10 mg/1 coumarine and 6 mg/1 dichloro-acetyl-hexa- methylene-imiπe. After 3-4 weeks the removal and planting can be repeated. Plants can be pinched back to keep the pr small size of plants. These cuttings without roots can be used for further propagation and tuberset. The "Somogy Gybngye" plants induced and grown in this mann 2 are producing 3500 pcs minitubers/m in 4 months.

Claims: Claims 1. A process for the production of potato minitubers from in vitro plants propagated by tissue culture and rooted in liquid media, characterized in that, tuber formation is in¬ duced in vitro on rooted plants, the induced plants are planted into solid media in glasshouses and tubers are de- .. veloping in glasshouses, while the growth of plant's haulm is limited by s praying with antigibberellic chemicals, regularly and periodically, the developed minitubers are collected, than plants are planted back into soil, the growth of haulm is limited further, and the removal and planting back are repeated periodically during the vegeta¬ tion period of plants. 2. A process as claimed in claim 1, characterized in that tuber formation is induced by using MS basic media contain¬ ing NH.NO- and KNO, at 1/10 concentration, 20 mg/1 glutamic acid, 40 g/1 saccharose, 5 mg/1 beπzyle adenine, 25 mg/1 coumarine and 6 mg/1 dichloro-acetyl-hexamethylene-imine. 3. A process as claimed in claim 1, characterized in that before planting into glasshouses plants are treated with a chemical which promotes rooting and with an agent whjch prevent the infection of Rhizoctonia, if needed. 4. A process as claimed in claim 3, characterized in that, this treatment is done by using indole-acetic acid and - if needed - propyl-N-(3-dimethyl-amino-propyl)-carbamate hydrochloride containing solution. 5. A process as claimed in any one of claims 1 to 3, charac¬ terized in that the transplanted plants are treated with coumarine and/or dichloro-acethyl-hexamethylene-imine. 6. A process as claimed in claim 1, characterized in that after collecting minitubers plants are treated again - befor transplantation - with a solution containing a chemical which promotes rooting and another one which prevent the infection of Rhizoctonia, if needed. 7. A process as claimed in claim 1, characterized in that the transplanted plants are treated with coumarine and/o dichloro-acetyl-hexamethylene-imine. 8. A process as claimed in any one of claims 1 to 7, characterized in that removal of plants, collection of minitubers and planting back the plants can be repeated several times during the vegetation period of plants.

Inventors: BALOGH JOZSEF, FOEGLEIN FERENC, OSVATH ZOLTAN, BALOGH, JOZSEF, FOEGLEIN, FERENC, OSVATH, ZOLTAN
Applicants: NOVOTRADE R T, NOVOTRADE RT.
Application year: 1987
Main classification: A01G7
IPC codes: A01H4, C12N5