Genetic Transformation By Non-tissue Culture On Phyllostachys Edulis And Phenotype Analysis Of The Offspring Of 35S::DhNiR Rice

There is a factor of the limiting about further enhance for bamboo industry,which is the difficulties of traditional hybridization of bamboo and selective breeding.It has resulted in lagging behind the breeding level.Rarely reports for the success of genetic transformation,result from the transgene is an effective breeding method,but difficulty for bamboo regenerated.There have been two aspects in this study,on the one hand,to avoid the problems about bamboo tissue culture through the research of the non-tissue culture.On the other hand,it is a research on genes that can promote plant regeneration,such as DhNiR(Dendrocalamus hamiltonii,Dh).To improve bamboo regeneration and transformation efficiency,we can by promoting the integration of regenerated genes into binary expression vectors.The purpose of this experiment is to apply two methods to bamboo,so that the transformation efficiency of bamboo has been effectively improved.The CP4-EPSPS gene was successfully transformed into bamboo seed by Agrobacterium tumefaciens-mediated plant germination seed gene transformation and the transgenic plants were successfully obtained.The results were as follows: the optimum concentration of bacteria was OD600 =0.07 in the co-culture.When the concentration is too low or too high,it will not conductive to the transformation and germination of seedlings.And the seedling rate could be increased by about 8%when adding 400 ?mol·L-1acetosyringone to co-culture medium.Finally,a strain of Phllostachys edulis transgenic plants was obtained by PCR.Followed by analysis of the offspring of 35:: DhNiR rice of phenotype.The test results are as follows: The experimental materials were the mature seeds of transgenic DhNiR and wild-type rice.After callus induction and proliferation,the callus of well-grown was selected for differentiation callus,there were obvious differences on the callus induction rate and the differentiation time.The frequency of green callus of DhNiR was 100%,while the rate of green callus of wild-type rice was only 56.37%when the callus was transferred to the differentiation medium on the 25 th day.And the seedling differentiation time is earlier than wild-type rice about 5 days.All of these results indicating that DhNiR gene can promot the regeneration ability of rice.The activity of nitrite reductase in each sample was determined by using the leaves,stem,root and callus of transgenic rice with DhNiR gene and wild-type rice.The activities of nitrite reductase in DhNiR rice were higher than those in wild-type rice,and the contents of nitrate and nitrite were studied respectively.The results showed that the contents of nitrate and nitrite in DhNiR gene were also higher than those in wild-type rice.