Study On In Viro Induction Of Autotetraploid Of Tomato (Lycopersicon Esculentum Mill.)

Tomato( Lycopersicon esculentum Mill.)is one of the important crops,which planted all over the world. Tomato breeding has got a lot of outstanding achievements due to numerous researches on it. To induce polyploid by manual way is one of important methods of tomato breeding, but it is seldom reported about how to induce autotetropolid of tomato by tissue culture. In this paper, in viro induction of autotetropolid tomato with colchicine was studied on basis of building regeneration system, the purpose of this paper is providing a new way for tomato variety improvement and germ-plasms source innovation.In the experiments, explants of tissue culture were cotyledons and hypocotyls from sterilitas tomato plants, results of research were as follows: the optimal way of seed sterilization was 10% NaClO +tween-20 2 drops for 30 min.The optimal culture medium for sterilitas plants was 1/2MS+sucrose10g/L + Carrageenan 7.5 g/L (pH5.8) ; the appropriate medium for cotyledon and hypocotyl explants differentiation is MS+ sucrose 30g/L+ Carrageenan 7.5 g/L+6-BA 1mg/L+ IAA 0.2 mg/L, with high differentiation rate, 96% and 91%, and 6.4 and 4.8 buds per explant respectively. The best medium for rootage was MS+ sucrose 20g/L+ Carrageenan 7.5 g/L+IAA 0.1 mg/L, with 100% rootage rate and good root system status.Two kinds of callus: undifferentiation callus and callus with buds were induced by the colchicine soaking method and colchicines medium method, the results showed: differentiation rate of callus declined sharply after dealing with colchicines, average differention rate of cotyledon callus was 60.9 %, 63.4% for comparison; the rate of hypocotyl callus is 48.6% and 53.7% for comparison. Several tetraploid plants were obtained by the colchicine soaking method and colchicines medium method, but undifferentiation callus were not fit to be used as induction material because of their low regeneration rate.Many autotetraploid plants were obtained by colchicines medium method to induce callus with buds. The cotyledon callus were cultured in medium with 0.2% colchicines for 48 and 72 hours, and the highest induction rate was 50%; the highest induction rate of hypocotyl callus was 50% with 0.1%, 0.2% and 0.4% colchicines for 48 hours; Soaking explants with colchicines restrained the growth of a great number of buds, and death rate was much higher than that of colchicine medium method. Two kinds of explants induced by different methods got different results, induction rate in turn was cotyledon and hypocotyl with colchicines medium method, then cotyledon and hypocotyl with colchicine soaking method, but results of death rate was reversed. The appropriate induction method of tetraploid tomato was culturing cotyledon and hypocotyl callus with buds in medium with 0.2% colchicines for 48 hour.Some indexes of diploid and tetraploid plant were compared and analysed, such as morphological character, stoma and physiological and biochemical indexes. Autotetraploids were stronger than diploidies cultured in the same media, with greener leaves and more roots, but their morphological character was similar. The prominent difference of morphological character was observed after transplanting for 40d. Tetraploid plants were much higher and stronger. Leaves length and wideth size increasing by 33.7% and 50% respectively, and leaf shape index got smaller. Guard cell size of tetraploid leaves was bigger than that of diploidies. Chloroplast number of gurad cell in tetraploids was 12.5, but only 8.6 in diploidies. In addition, the change of stoma size was similar with gurd cell size, but stoma density in tetraploids diminished significantly.Contents of chlorophyll and carotenoid of tetrapliod leaves were higher than those of diploidies, but there was not significant difference between ratio of chlorophyll a and b in different ploidy plants. MDA content of tetrapliods was lower than of diploids, but POD activity and SOD activity and soluble protein content of tetrapliods were higher. By comparison, the physiological status of tetraploids was more excellernt in the same growth environment. The result of peroxidase isozyme analysis was as follows: the peroxidase isozyme band of tetraploids had more two bands of A and F besides eight same bands, at the same time the colar of bands was enhanced. The peroxidase isozyme analysis was useful for indirect judgement of ploidy tomato by isozyme analysis.