Objective: In the design of recombinant protein expression conditions, a number of essential elements such
as type of medium and bioprocess parameters like temperature, agitation, dissolved oxygen, trace elements
optimization is considered. As bioprocess is cost effective process, hence production reproducibility issues
can be addressed by well designed, medium and protein expression influencing parameters. In this study,
rDNA E. coli chosen as protein expressive organism. To maximize the expression of protein different types
of media and trace elements, temperature and RPM conditions were studied. This study goal was to develop
optimized media to maximum expression of protein (Cross reactive Material (CRM197) of Diphtheriae
toxoid) in recombinant E. coli (E. coli Strain-BL21 DE3, Plasmid: PET24a), which is used for conjugation
of the polysaccharide vaccines to produce the T dependent immune system in children. Materials and
Methods: rDNA E. coli, LB medium, Trace elements, Probe Sonicator and refrigerated centrifuge. For
growth of the rDNA E. coli, three different media (Minimal, Medium and Rich Media) were selected, and
temperature 25 ºC,37 ºC, RPMs were selected to observe expression of protein. The expression of CRM197
was analyzed by using the SDS-PAGE Gel electrophoresis. Lysis and formation of IBs were purified to get
purified protein. Conclusion: The result revealed that the Rich media and Trace Element Zinc sulphate
combination holds potential for use as a high cell density growth medium and to get high intensity of the protein expression CRM197 for recombinant E. coli (CRM197), when compared with Minimal Medium.
Keywords: CRM197, Diphtheriae toxoid, Rich Media, SDS PAGE, recombinant E. coli,
polysaccharide vaccine
Publication date: 15/06/2023
https://ijbpas.com/pdf/2023/June/MS_IJBPAS_2023_JUNE_SPCL_10451.pdf
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https://doi.org/10.31032/IJBPAS/2023/12.6.1045
