The Approach of High-Efficiency Production of Recombinant Protein in Rice Suspension Cells- Alternative Carbon Source and a Novel Immobilization Bioreactor

Recent developments in tissue culture techniques make it possible to use transgenic plant cells to produce valuable heterogonous proteins under controlled conditions, offering the prospects of high productivity and great flexibility in production of pharmaceutical proteins. Plant-suspension cell system can produce recombinant proteins in large scale with high efficiency in bioreactor. Therefore, development of scale-up production technology of high-value proteins by plant suspension cells system is the important issue in plant biotechnology fields. Rice αAmy promoter is a potential alternative for high production of valuable heterogonous proteins under inducible condition. However, medium exchange to deplete sugar in culture medium may induce cell death due to the loss of carbon energy sources in 91/10/30 修訂 metabolism. Therefore, addition of some alternative carbon sources have been applied to improve recombinant protein production. In this study, we explored ways to improve the production yield of recombinant stable recombinant human epidermal growth factor fusion protein (H-EGF) using some alternative carbon sources for recombinant H-EGF production using modified αAmy8 promoter and αAmy3 signal peptide in large-scale culture. Immobilized plant cell cultures have a number of advantages over suspension cells in the practical field of bioreactor and in the physiological state of the cells during the culture period. Also, immobilization offers an easily manipulated low shear environment. In this study, an airlift-stirred combination reactor is developed and designed for the continuous production of recombinant H-EGF. The airlift-stirred combination reactor（ASCR） not only adjusts separately, but also proceeds simultaneously mixing and aeration under various operating conditions. Therefore, it is expected that the ASCR provides high oxygen transfer rates and maintains the immobilized cells in a low-shear environment. In this study, we will develop a simple and rapid technology for efficient heterologous protein production in immobilized transgenic rice cell cultures. Here, five keystone technology will be developed：（1）The optimum cultivation strategies using modified αAmy8 promoter in medium containing the alternative carbon source；（2）The investigation of optimum cultivation medium and additives；（3）A scale-up immobilization fermentation process for efficient target proteins production；（4）A novel ASCR bioreactor development and evaluation for high-yield H-EGF production；（5）The establishment and evaluation of a simple and rapid method for the purification of H-EGF from bioreactor.

Project ID:PA10507-1179
External Project ID:MOST105-2311-B182-002