The Establishment Of Tissue Culture System Of Saintpaulia Ionatha ’Hiroko’ For The Purpose Of Agrobacterium-mediated Transformation

This study took African violet (Saintpaulia ionatha)’Hiroko’as experimental material,through adventitious buds induction, adventitious shoots proliferation and plantlets ex vitro rooting to create a tissue culture system of African violet. Meanwhile, on the basis of the tissue culture system, we studied the effects of different antibiotics on the adventitious shoot regeneration and adventitious roots induction of African violets, with a view to provide a basis to subsequent genetic transformation of African violets.The tissue culture systems establishment of African violet:6-BA/NAA, sucrose concentration and different site of leaves were chosen as a test factor through orthogonal experiment to determine the most suitable conditions for adventitious buds induction at the adventitious buds induction stage, the most appropriate explants were the lower part of the leaves, and the best culture medium was MS+6-BA (0.5mg/L)+NAA (0.5mg/L)+sucrose (20g/L). At the adventitious shoots proliferation stage,6-BA/NAA, GA concentration and sucrose concentration in the medium were chosen as a test factor through orthogonal experiment to determine the most suitable conditions for adventitious shoots proliferation, and the best culture medium was MS+6-BA (0.1mg/L)+NAA (0.1mg/L)+GA (2mg/L)+sucrose (20g/L). The most suitable conditions for their rooting was selected through the single factor cottage test on the plantlets in different growth conditions, at the plantlets ex vitro rooting stage. The first-class seedlings (grown to2cm, more than three pairs of mature leaves, robust growth without deformity) can be rooted directly into the matrix, and the most appropriated cutting medium was vermiculite. The second-class seedlings (grown to2cm, more than two pairs of mature leaves, robust growth without deformity) were best to start with a quick dip into200mg/L of NAA before cutting to the vermiculite and after doing this the rooting rate can reach100%.Effects of several antibiotics on adventitious shoot regeneration and adventitious roots induction of African violet:we studied the effects of three antibacterial antibiotics (cefotaxime, cephalexin, carbenicillin) on adventitious shoot regeneration of African violet, and the effects of three selective antibiotics (kanamycin sulfate, Neomycin sulphate, G418) on adventitious shoot regeneration and adventitious roots induction of African violet. According to the results, the most suitable antibacterial antibiotic used in shoot regeneration stage were carbenicillin and cefotaxime, and the optimum concentrations were both300mg/L, while cephalexin was not a preferred choice. The most suitable selective antibiotic used in shoot regeneration stage were kanamycin sulfate and G418, and the optimum concentrations were50mg/L and5mg/L, while Neomycin Sulphate was not a preferred choice. The most suitable selective antibiotic used in adventitious root induction stage were kanamycin sulfate, and the optimum concentrations was80mg/L, while Neomycin Sulphate and G418were not suitable.