Munjal, Neera (2014-12). Bioprocessing of Microalgae C. reinhardtii for Production and Purification of Single Chain Antibody Fragment. Master's Thesis.
Thesis
The eukaryotic green alga, Chlamydomonas reinhardtii, is a unique expression platform that can efficiently express complex therapeutic proteins. However, demonstrating that therapeutic molecules can be produced in quantifiable levels is essential to establish the potential of the C. reinhardtii expression system. Thus, the objective of the first part of investigation (Chapter II) was to determine the process conditions that could maximize C. reinhardtii biomass accumulation and induced-production of the two recombinant proteins, a single chain fragment antibody molecule (?CD22 scFv) and malaria vaccine antigen (Pfs25), produced in the chloroplast of C. reinhardtii. To achieve a higher production of recombinant proteins, cultivation variables of C. reinhardtii, such as mixing, light-induction time and intensity, nutrient depletion and culture age, were investigated and optimized. The optimal light-induction time was 24 h at a light intensity of 300 umol m^-2 s^-1. Replacement of the culture media in the late exponential growth with fresh media was beneficial to the accumulation of recombinant proteins. Optimization led to increases in the accumulation of recombinant proteins by six-fold and the recombinant protein fraction in the extracted soluble protein by two-fold.
