Establishment And Optimization The Transgenic System Of Avena Nuda L.

In this experiment,mature embryos of eight naked oat varieties were used as explants,and the dedifferentiation and redifferentiation of mature embryos of naked oats were explored through the study of varieties,medium,hormone types and concentration ratios in callus culture.Influencing factors in the process,and select varieties with high regeneration frequency;Using Agrobacterium-mediated transformation of wounded embryos,by studying the concentration of antibiotics in the medium,the concentration of Agrobacterium infection and the transplanting time,the best conditions for wounded embryo culture are obtained,and a rapid genetic transformation system for wounded embryos is established.The results are as follows:1.There is an interaction relationship between varieties and culture medium.Different varieties of oat mature embryos have different healing rates in the same culture medium,and there are also big differences in the healing rate of the same varieties in different media."Bayou No.1" is the most suitable for callus induction on MS medium,with a callus rate of 87.33% and a plant regeneration rate of 32.99%,which can be used for the establishment of a naked oat regeneration system and further genetic transformation.2.The best medium for callus culture of naked oat variety "Bayou No.1" is as follows:Callus induction medium MS+2.0 mg/L 2,4-D+1.0 mg/L NAA+1.0 mg/L IAA+500 mg/L hydrolyzed casein(CH)+25 g/L sucrose;callus subculture medium:MS+1.5 mg/L 2,4-D+1.0 mg/L NAA+1.0 mg/L 6-BA+ 30 g/L sucrose;Callus Green Bud Differentiation Medium: MS+1.5 mg/L 6-BA+0.5 mg/L NAA+0.5 mg/L IAA;callus rooting medium: 1/2 MS+0.5 mg/L NAA+0.1% rooting powder.3.Using "Bayou No.1" as the material,the optimal concentration of the bacteriostatic ampicillin in the culture medium during the process of Agrobacterium transformation of wounded embryos is 200 mg/L,and the optimal concentration of the selective agent kanamycin is 40 mg/L.4.Use different concentrations of Agrobacterium to infect "Bayou No.1" wounded embryos,the optimum concentration of OD600 is 0.8.5.Wounded embryos cultured in the culture medium for different time affects the survival rate.After the transformation,it is best to cultivate in the medium for 10 days.6.The plasmid pBI121-GFP was transformed into "Bayou No.1" wounded embryos.After screening and culture,126 regenerated plants were obtained.PCR detection was performed on the regenerated plants,and 4 positive plants were obtained.The transformation rate of naked oat wounded embryos was 3.17%.