In this thesis,the best induction method of polyploid from grape shoots under field callus and tissue culture conditions was studied.The effects of different wrapping methods,hormone concentration,shoot thickness,short cut and genotype on callus induction of grape new shoot were investigated under field conditions,and colchicine treatment was applied to callus of grape new shoot.Under tissue culture conditions,the effects of different treatments on seed germination rate and colchicine on induction efficiency of’Rose fragrance’and’Jintian Royal seedless grape’were studied.At the same time,polyploid mutagenesis treatment was carried out on tissue culture seedlings of’Jintian Royal seedless’grape.The results are as follows:1.Callus induction of grape shoots in field.In July of the growing season,the new shoots of grape with a diameter of more than 1 cm were cut off between nodes,and the cross section was added with hormone TDZ 10.0 mg/L+Ag NO32.0 mg/L,and the triple long-term moisturizing method of black bag+wet mud+black bag was applied to cover and wrap.The callus induction rate of grape new shoots could reach 90%.This study can provide reference for polyploid induction from grape callus in field.2.Seed germination of’Muscat Hamburg’.The seeds of"Muscat Hamburg"grape were soaked in 2500 mg/LGA3 for 48 h at room temperature.The beak-shearing treatment was carried out during inoculation,and the beak-shearing treatment was adopted at the time of inoculation.The germination rate of grape seeds was up to 84%when cultured in 1/2 ms+6%sucrose+0.6%agar medium.3.Seed colchicine mutagenesis.Colchicine mutagenesis was performed at different stages of seed germination.When radicle germinated to 0.5-1.0cm,colchicine medium with different concentrations was inoculated,colchicine medium with 40 mg/L was incubated,and then transferred to normal medium after 2 weeks of treatment.The phenotype variation rate was 15.55%.The mutant material was identified as tetraploid’Muscat Hamburg’by flow cytometry.4.Mutagenesis of tissue culture seedlings.One stem segment was inoculated with colchicine medium with a concentration of 60 mg/L by tissue culture of’Jintian Royal seedless’.After 2 weeks of treatment,the stem segment was transferred to normal medium,and the phenotypic variation rate was 10.00%.The mutant material was identified as tetraploid’Jintian Royal seedless’by flow cytometry.5.Identification of mutant materials.After mutagenesis,the leaves were larger,darker and shrunked,the leaf edges were long serrated,and the roots were thick.The stomatal length and width of tetraploid plants were larger than that of diploid plants.The DNA content of the mutant material was determined by flow cytometry,and the result was tetraploid.In this experiment,the tetraploid’Muscat Hamburg’and the tetraploid’Jintian Royal seedless’were obtained successfully.To obtain large grain’Muscat Hamburg’and large seedless’Jintian Royal seedless’to lay the foundation. |