A method to simplify bioreactor processing for recombinant protein production in rice cell suspension cultures

Abstract

Transgenic plant cell suspension culture is a promising platform for recombinant protein production. Rice cell suspension culture is one of the systems that has been developed due to its unique metabolically-regulated promoter, rice alpha-amylase 3D ( RAmy3D ), that is up-regulated in sugar-deprived medium. Using the RAmy3D promoter system in transgenic rice cell suspensions results in two phases of the culture, the growth phase and the induction phase. Conventionally, medium exchange is performed to remove residual sugar and induce recombinant protein. In this work, a simplified production process is demonstrated in a 5-L bioreactor, including reduction of sugar concentration in the initial culture medium, elimination of the media exchange operation, and uncontrolled dissolved oxygen (DO) with constant aeration. The simplified method significantly improves the accumulation level of a recombinant protein, protein purity, and productivity compared to the conventional method. This method also reduces costs associated with material and labor. • The method of simplified bioreactor processing includes single-stage culture, uncontrolled dissolved oxygen (DO) but controlled inlet air flowrate, and lower (50% reduction) initial sucrose concentration in the culture medium. • This method improves recombinant protein production level and productivity compared to the conventional method. • This method reduces material and labor costs.

Production of recombinant butyrylcholinesterase from transgenic rice cell suspension cultures in a pilot‐scale bioreactor

“…The inoculation corresponded to 15%–18% v/v (volume of inoculating culture to the final working volume of culture). Single‐stage operations were used in this study as previously described (Macharoen et al, 2020a, 2020b). The first bioreactor run was operated in a cyclical semicontinuous mode where the media exchange to start a new cycle was performed at Day 4 since extracellular sugar depletion by cellular uptake.…”

“…A recent study by our group using technoeconomic analysis showed that the cost of production and purification of rice-made rBChE (rrBChE) was dramatically reduced from the estimated $20,000 for a 400-mg dose from blood plasma to less than $300/dose of pure rrBChE using semicontinuous transgenic rice cell suspensions (Corbin et al, 2020). Here, we demonstrate the use of a 40-L conventional stainless-steel stirred tank bioreactor (STB) for scaling-up the production of rrBChE in transgenic rice cell suspension cultures under single-stage operations (rice cells naturally deplete sugar in the culture medium without medium exchange) in uncontrolled dissolved oxygen (DO) condition at a constant aeration rate (Macharoen et al, 2020a(Macharoen et al, , 2020b.…”