Studies On Polyploid Induction And Effect Of LED On Tissue Cultured Seedling In Stevia Rebaudiana(Bertoni) Hemsl.

Stevia rebaudiana(Bertoni)Hemsl.is a perennial herb of compositae and its leaves are rich in stevia glycosides.Which is the sweet raw material of foods and pharmaceuticals industries,and is internationally known as "the world's third healthy sugar source".In this study,the adventitious buds of stevia were used as experimental materials to establish the sterile rapid propagation of Stevia;On this basis,the effects of colchicine and trifluralin on stevia polyploid induction were discussed,the variant plants were identified and the content of stevia glycosides in the obtained polyploid plants was determined;The effect of LED light quality on proliferation,physiological and biochemical indexes of tissue culture seedlings was studied to obtain the best LED light quality for vitro culture of stevia.Providing important scientific evidences for improving the reproduction coefficient of stevia,breeding new varieties with high stevia glycoside content and excellent characters,carrying out factory scale production and meeting the needs of market.The main results are as follows:1.The regeneration system was established with stevia stem segments with axillary buds as explants.The best method of disinfection for explants was 70%alcohol disinfection for 30 s and 0.2%sodium hypochlorite treatment for 10 min;the optimal medium for adventitious bud proliferation was MS+0.5 mg/L 6-BA+0.1 mg/L NAA;and the best medium for seedling and rooting was MS+0.5 mg/L IBA+0.5 mg/LNAA.2.The effects of colchicine and trifluralin on the induction of autotetraploid in the adventitious buds of stevia were studied.The adventitious buds were soaked with 0.1%colchicine for 24 h,and the induction rate was 23.1%.When the concentration of colchicines was 100 mg/L with 21 d treated time,the induction rate reached 27.5%.The adventitious buds were soaked in 30 mg/L trifluralin treatment for 48 h to the best induction effect,and the induction rate reached 33.2%.3.Morphological observation,cytological identification and chromosome count were used to identify the variant plants.The leaves of tetraploid plants showed obvious enlargement and partial shrinkage The stomatal length and width of tetraploid leaves were 160.78%and 154.15%of those of diploid leaves respectively.The number of stomata in tetraploid leaves under the same visual field(eyepiece 10 times,objective 40 times)was only 48.17%of that of diploid leaves.The number of chromosomes in stevia diploid control plants was 2n=2X=22,and that in tetraploid plants was 2n=4X=44.4.The content of stevia glycosides in the leaves of tetraploid plants was determined.The total content of stevia glycosides in the leaves of tetraploid plants of "nongda l" was 4.16%higher than that of diploid plants,the content of RA glycosides was 6.98%higher,and the content of RM glycosides was 0.51%higher.The total glycosides in the leaves of tetraploid plants of "nongda3"was 2.88%higher than that of diploid plants,the content of RA glycosides is 9.38%higher,and the content of RM glycosides was 0.66%higher.5.When LED red light:blue light=3:1,was suitable for adventitious bud proliferation and growth of ground parts and chlorophyll content accumulation,adventitious bud proliferation multiple was 13.9,relatively strong;Plants height was 82.57 mm.The fresh weight and dry weight on the ground reached the maximum of 1.95 g and 0.16 g respectively.The total chlorophyll content reached the maximum of 1.98 mg/g.The number of roots increased under the treatment of monochromatic light quality and combined light quality.Under the treatment of single red light,the number of roots reached the maximum of 16.4,but LED light quality inhibited root elongation and root fresh weight and dry weight.When red light:blue light=1:2,the fresh weight and dry weight of the whole plant were the highest,which were 3.68 g and 0.29 g respectively.