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Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011elements that regulate plant growth and development. The form of nitrogen supplied toplants may influence the uptake of other nutrients (Schmitz and Lorz, 1990). A low level ofammonium supplied in combination with nitrate is favorable to tuber growth, but a higherlevel of ammonium supply is detrimental (Garner and Blake, 1989). In MS basal saltmedium the two main sources of nitrogen (N): ammonium nitrate (NH 4 NO 3 ) and potassiumnitrate (KNO 3 ).The present experiment was undertaken to study the effect of inorganic nitrogensupply on the induction and development of potato microtubers “in vitro”.2. MATERIALS AND METHODSNicoleta and Christian potato (Solanum tuberosum L.) varieties were used in theexperiment.“In vitro” plantlets of potato were multiplied by subculturing of single stem nodes.The multiplication medium contained mineral salts and vitamins plus 0.05 mg/l NAA and20g/l sucrose. The medium was solidified with 8 g/l Agar. Temperature in the growthchamber was 25 o C with 16 hr photoperiod and light by fluorescent tubes.Fifteen explants (each with two nodes) were cultured for each treatment andcultivar, in vessels. The microtuberization medium is liquid and contains the samesubstances as medium Murashige-Shook (1962) coumarin, kinetin, sucrose (80-90 g/l). Thestock solution of cumarine and kinetin were prepared as follows:Cumarine: 500 mg in a 200 ml recipient; 25 ml ethanol are added; the solution iscompleted with distilled water.Kinetin: 200 mg in a 50 ml recipient; 20 ml NaOH 1N are added; the solution iscompleted with distilled water.For 1 l medium we add 20 ml stock solution of coumarine and 12.5 ml stocksolution of kinetin. The microtuberisation medium doesn’t contain agar. In every plasticrecipient we poured 45 ml medium The microtuber induction cultures were incubated in thedark at 20 0 C. We harvested the microtubers separately according to variety.After the tuberization period (7-8 weeks of darkness), plantlets were extractedfrom cultures recipients, and harvested microtubers were washed, to avoid the subsequentinfections, which can appear during their storage. The tuberization percentage (number ofexplants with at least one microtuber over the total number of explants cultured) wasmeasured.In nitrogen-rich MS medium the concentration of total inorganic nitrogen is 60meq. In the present experiment, alterations in MS macronutrients were made to obtain threelevels of total nitrogen (30, 45 and 60 meq) while keeping potassium level constant at 20meq. The different media used in the experiment are shown in Table 1.Observations were recorded on the number of microtubers per g vessel, averagemicrotuber weight.3. RESULTS AND DISCUSSIONReducing the total nitrogen supply increased the number but decreased the size ofmicrotubers. Reduced total availability of inorganic nitrogen in the medium caused a52
Proceeding of the 4 rd International Symposium“NEW RESEARCH IN BIOTECHNOLOGY” USAMV Bucharest, Romania, 2011decline in the average microtuber weight or size of the microtubers, suggesting that themineral nitrogen nutrition is a major limiting factor controlling their size.Table 1. Total of nitrogen used in nutrition mediumNitrogenSalts(meq) KNO NH NO3 4 3(mg l -1 ) mM (mg l -1 ) mM30 950 9.4 825 10.345 1425 14.1 1237 15.560 1900 18.8 1650 20.6Higher numbers of microtubers could be obtained using MS mediumsupplemented with 30 meq total nitrogen, 20 meq potassium, 80 g 1 -1 sucrose, but increasein microtuber numbers was associated with simultaneous decrease in microtuber size. Forcommercial seed potato production, microtuber size is more important than microtubernumber because small sized microtubers are more vulnerable to storage loss (Naik &Sarkar, 1997) and are unsuitable for direct field planting (Jones, 1988).Table 2. Influence of nitrogen concentration and on weight of microtubers produced/ microplantVariety Total nitrogen Average weight of a % Dif. (g) Signif.(meq)microtuber (g)60 1.68 100.00 -NicoletaChristian45 1.25 74.91 -0.42 ns30 1.01 60.46 -0.66 o60 1.91 100.00 -45 1.49 78.50 -0.41 ns30 1.14 60.09 -0.77 oLSD 5%= 0.60 g LSD 1%=0.80 g LSD 0.1%=1.03 gns= not significantThe weight of microtubers per vessel was the highest at the highest nitrogenconcentration (60 meq) (table 2): 1.91 g for Christian variety and 1.68 g for Nicoletavariety. The lowest weight of microtubers was obtained at the lowest nitrogen level 30meq: 1.14 g for Christian variety, with a difference of -0.77 g, significant in a negative way,and 1.01 g for Nicoleta variety, with a difference of -0.66 g, significant in a negative way.The number of microtubers per vessel was the highest at the lowest nitrogenconcentration (30 meq) (table 3): 19.67 microtubers, with a difference of 4.34 microtubers,significant in a positive way, for Christian variety, followed by Nicoleta with 18.33microtubers and a difference of 3.00 microtubers, significant in a positive way.The number of microtubers per vessel was the lowest at the highest nitrogenconcentration (60 meq): 15.33 microtubers, for both varieties.53
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