- 學位論文
Development of Tissue Culture Techniques to Improve Propagation in Saussurea involucrata and Secretory Protein Production in Oryza sativa
Development of Tissue Culture Techniques to Improve Propagation in Saussurea involucrata and Secretory Protein Production in Oryza sativa
摘要
Plant tissue culture utilizes appropriate culture medium to propagate cells and tissue in sterile condition. This thesis combines two pilot studies regarding the use of the plant tissue culture techniques, one in Saussurea involucrata and the other in Oryza sativa. The first research is to speed-up the propagation process in S. involucrata. The method developed here was to establish S.involucrata suspension cells from leaf directly. To speed-up regeneration process in S.involucrata, the establishments of optimal medium such as combinations of plant hormones, proteins and nature ingredient into suspension culture medium were applied. The result showed that whole plant propagation took 104 days from embryonic cell to plantlets of S.involucrata, a month shorter than current record. Syringin, rutin, and hispidulin have been detected in callus, but hispidulin was not detected in suspension cultured S.involucrata. The second pilot study is to improve secretory protein production in Oryza sativa. Thirty percent increases of total secretory proteins in rice suspension culture can be achieved by modification of MS medium. The supplement of nature ingredient also gave the best results with 32% increase of total proteins. Elimination of 2, 4-D, approximately 40 KDa of alpha-amylase proteins was observed in culture medium by silver staining, implying that alpha-amylase promoter driving recombinant protein system can be expressed without 2,4-D.
並列摘要
Plant tissue culture utilizes appropriate culture medium to propagate cells and tissue in sterile condition. This thesis combines two pilot studies regarding the use of the plant tissue culture techniques, one in Saussurea involucrata and the other in Oryza sativa. The first research is to speed-up the propagation process in S. involucrata. The method developed here was to establish S.involucrata suspension cells from leaf directly. To speed-up regeneration process in S.involucrata, the establishments of optimal medium such as combinations of plant hormones, proteins and nature ingredient into suspension culture medium were applied. The result showed that whole plant propagation took 104 days from embryonic cell to plantlets of S.involucrata, a month shorter than current record. Syringin, rutin, and hispidulin have been detected in callus, but hispidulin was not detected in suspension cultured S.involucrata. The second pilot study is to improve secretory protein production in Oryza sativa. Thirty percent increases of total secretory proteins in rice suspension culture can be achieved by modification of MS medium. The supplement of nature ingredient also gave the best results with 32% increase of total proteins. Elimination of 2, 4-D, approximately 40 KDa of alpha-amylase proteins was observed in culture medium by silver staining, implying that alpha-amylase promoter driving recombinant protein system can be expressed without 2,4-D.