Study On Rapid Propagation In Vitro And Polyploid Induction Of ’ Lv Ling ’ Walnut

In order to establish the tissue culture and rapid propagation system of walnut and establish the polyploid breeding technique system of walnut, the ’Lv Ling’ walnut seeds, the annual ’Lv Ling’ walnut seedling shoots and 5 years old ’Lv Ling’ walnut scion nursery shoots were used as materials, the rapid propagation of ’Lv Ling’ walnut seedlings by embryo axillary bud without leaf and the rapid propagation of axillary bud were studied, polyploid plants of ’Lv Ling’ walnut were inducted with the methods of shoot apex infiltration, seedlings top-pinching infiltration, shoot top-pinching infiltration, seeds kernel immersion, embryo immersion, kryptoblast stimulation and timulating axillary bud stimulation, the variant stains were identified by flow cytometry. The results were as follows:1. The germination induction medium of rapid propagation of axillary bud without leaf was MS + IBA 1.0 mg/L + 6-BA 1.0 mg/L + sucrose 30 g/L + agar 6 g/L, the germination rate was 100%, the multiplication coefficient was 5.84. The elongation medium of seed embryo axillary bud without leaf was MS + 6-BA 2.0 mg/L + IAA 1.0 mg/L + GA3 1.5 mg/L+ sucrose 30 g/L + agar 6 g/L,and in the medium, elongation of axillary bud without leaf was longest, and its growth condition was the best; the germination medium of embryo in the rapid propagation of axillary bud was MS + sucrose 30 g/L + agar 6 g/L, the germination rate and seedling rate were 90%, the germination medium of axillary buds was MS + 6-BA 2.0 mg/L + NAA 0.2 mg/L + sucrose 30 g/L + agar 6 g/L, the germination rate of axillary buds and multiplication coefficient were 80% and 3.54.2. Two-stage rooting method was used to induced walnut rooting that 2 mm digestion stem segment from petiole base was cultivated in DKW + IBA 8.0 mg/L + sucrose 30 g/L + agar 6.0 g/L for 20 days under dark conditions firstly, then transferred in DKW+ sucrose 30 g/L + agar 6.0 g/L under light condition, the rooting rate was 50%. Tissue culture seedlings of induced rooting were transplanted to field with the survival rate 33.3%.3. The variation rates were 0% respectively with the methods of shoot apex infiltration, shoot top-pinching infiltration, seeds kernel immersion, kryptoblast stimulation and timulating axillary bud stimulation; with the method of seedlings top-pinching infiltration, there were 4 varied plants in 30 seedlings after infiltrated with 1.0% colchicine solution for 8 d, and the variation rate was 13.3%. The plants whose leaves were notably larger, thicker, darker and wrinkled than that of normal were identified, the result showed that1plant was tetraploid, 3 plants were mixoploidy. The walnut embryos stripped after harvested in the kernel enrich stage were immersed in different colchicines solution, the variation rate was 23.3% by 0.4% colchicines solution for 12 h, 1 plant was tetraploid; the variation rate was 20% by 0.8% colchicines solution for 12 h, 1 plant was tetraploid; the variation rate was 50% by 0.4% colchicines solution for 24 h, 2 plant was tetraploid, 1 plant was mixoploidy.