Propagation of plant cells in robust single-use bioreactors

Abstract

New and effective technologies are needed to full fill the increasing demands of sustainable food production due to the growing population, urbanization, climate change, and decreasing water supplies. Plant cell cultivation in closed bioreactor systems offers an attractive new approach for this challenge. Innovation about the production of plant cells in the non-laboratory environment has been studied in VTT Technical Research Centre of Finland. This thesis continued the development of home bioreactor for the robust propagation of plant cells as a food source. The first aim of this study was to find out which are the key physical factors affecting the biomass accumulation of lingonberry cell suspension culture. Multiple growing experiments under shake flask scale and 2 L working volume stirred tank cultivations were carried out to get more detailed data about growth parameters of lingonberry cell culture. For effective growth without long lag phase inoculum density should be above 60 g/L. Lingonberry cell’s need of dissolved oxygen is surprisingly moderate, optimal target would be 5-10 %. To maintain sufficient gas exchange level and prevent cell sedimentation, gentle but effective mixing is needed. The light had a most remarkable impact on biomass accumulation. Cells cultivated in dark produced 47 % more fresh biomass compared to cells grown under white light with 16:8 h photoperiod. Pigment formation revealed to be reversible and dependent on sufficient amount of light. All previous trials for lingonberry cell cultivations in simple single-use bags have been failed. Gained new information about physical growth demands from this study made designing of suitable single-use bioreactor now possible. Based on the results optimal and as simple as possible single-use bioreactor prototype and cultivation bag were designed. The bag in a box -solution with orbital shaking was chosen. In this model mixing and aeration can be introduced in a simple way without sparging and risk of foaming. Lingonberry cells were successfully cultivated in single-use cultivation bag and biomass accumulation reached promising levels (174 g/L) compared to shake flask (208 g/L) and stirred tank bioreactor cultivations (210 g/L). Based on all the results of this study, lingonberry cell suspension cultures can now be cultivated in a simple single-use bioreactor. Development of robust home usable bioreactor for plant cell production can be continued and future decisions related to prototype design and building can be based on solid data. However, to achieve optimal biomass accumulation, more studies related to process optimization, especially in lighting and mixing conditions, should be carried out with the designed single-use bioreactor