Rapid Strain Evaluation Using Dynamic DO-Stat Fed-Batch Fermentation Under Scale-Down Conditions

Part of the Methods in Molecular Biology book series (MIMB, volume 834)


While large amount of strains can be quickly generated via metabolic engineering, the speed/efficiency of evaluating each strain becomes the bottleneck in the process from strain development to final production. In this chapter, a method is introduced to rapidly evaluate strain performance in fed-batch fermentation mode by using dynamic dissolved oxygen stat feed back control with no additional advanced online measurement. In addition, a scale-down feature is integrated in the method to mimic the limitation of oxygen transfer in large-scale vessels, so that strains can be evaluated under the conditions close to that in large-scale bioreactors. The method has been implemented in several commercial standard benchtop scale fermentation systems with different fermentation control software.

Key words

Strain evaluation Fed-batch fermentation Dynamic DO-Stat control Scale-down Process development 


  1. 1.
    Warner, J. R., Reeder, P. J., Karimpour-Fard, A., Woodruff, L. B. A., and Gill, R. T. (2010) Rapid profiling of a microbial genome using mixtures of barcoded oligonucleotides, Nat Biotechnol 28, 856–862.PubMedCrossRefGoogle Scholar
  2. 2.
    Gill, R. T., Wildt, S., Yang, Y. T., Ziesman, S., and Stephanopoulos, G. (2002) Genome-wide screening for trait conferring genes using DNA microarrays, Proc Natl Acad Sci USA 99, 7033–7038.PubMedCrossRefGoogle Scholar
  3. 3.
    Isett, K., George, H., Herber, W., and Amanullah, A. (2007) Twenty-four-well plate miniature bioreactor high-throughput system: Assessment for microbial cultivations, Biotechnol Bioeng 98, 1017–1028.PubMedCrossRefGoogle Scholar
  4. 4.
    Huber, R., Ritter, D., Hering, T., Hillmer, A. K., Kensy, F., Müller, C., Wang, L., and Büchs, J. (2009) Robo-Lector – a novel platform for automated high-throughput cultivations in microtiter plates with high information content, Microb Cell Fact 8, 42.PubMedCrossRefGoogle Scholar
  5. 5.
    Tang, Y. J., Laidlaw, D., Gani, K., and Keasling, J. D. (2006) Evaluation of the effects of various culture conditions on Cr(VI) reduction by Shewanella oneidensis MR-1 in a novel high-throughput mini-bioreactor, Biotechnol Bioeng 95, 176–184.PubMedCrossRefGoogle Scholar
  6. 6.
    Kensy, F., Engelbrecht, C., and Büchs, J. (2009) Scale-up from microtiter plate to laboratory fermenter: evaluation by online monitoring techniques of growth and protein expression in Escherichia coli and Hansenula polymorpha fermentations, Microb Cell Fact 8, 68.PubMedCrossRefGoogle Scholar
  7. 7.
    Ingo Knabben, Regestein, L., Marquering, F., Steinbusch, S., Lara, A. R., and Büchs, J. (2010) High cell-density processes in batch mode of a genetically engineered Escherichia coli strain with minimized overflow metabolism using a pressurized bioreactor, J Biotech 150, 73–79.CrossRefGoogle Scholar
  8. 8.
    Lara, A. R., Caspeta, L., Gosset, G., Bolívar, F., and Ramírez, O. T. (2008) Utility of an Escherichia coli strain engineered in the substrate uptake system for improved culture performance at high glucose and cell concentrations: An alternative to fed-batch cultures, Biotechnol Bioeng 99, 893–901.PubMedCrossRefGoogle Scholar
  9. 9.
    Lee, S. Y. (1996) High cell-density culture of Escherichia coli, in Trends in Biotechnology, pp 98–105. Elsevier Science Ltd.Google Scholar
  10. 10.
    Åkesson, M., Hagander, P., and Axelsson, J. P. (2001) Avoiding acetate accumulation in Escherichia coli cultures using feedback control of glucose feeding, Biotechnol Bioeng 73, 223–230.PubMedCrossRefGoogle Scholar
  11. 11.
    Åkesson, M., Karlsson, E. N., Hagander, P., Axelsson, J. P., and Tocaj, A. (1999) On-line detection of acetate formation in Escherichia coli cultures using dissolved oxygen responses to feed transients, Biotechnol Bioeng 64, 590–598.PubMedCrossRefGoogle Scholar
  12. 12.
    Luli, G. W., and Strohl, W. R. (1990) Comparison of growth, acetate production, and acetate inhibition of Escherichia coli strains in batch and fed-batch fermentations., App Environ Microbiol 56, 1004.Google Scholar
  13. 13.
    Mey, M., Maeseneire, S., Soetaert, W., and Vandamme, E. (2007) Minimizing acetate formation in E. coli fermentations, J Ind Microbiol Biotechnol 34, 689–700.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.DuPont Central Research and DevelopmentWilmingtonUSA

Personalised recommendations