Increasing the buffering capacity of minimal media leads to higher protein yield

Stephan B. Azatian, Navneet Kaur, Michael P. Latham

Research output: Contribution to journalArticlepeer-review

35 Scopus citations


We describe a general and simple modification to the standard M9 minimal medium recipe that leads to an approximate twofold increase in the yield of heterologously expressed proteins in Escherichia coli BL21(DE3) bacteria. We monitored the growth of bacteria transformed with plasmids for three different test proteins in five minimal media with different concentrations of buffering salts and/or initial media pH. After purification of the over-expressed proteins, we found a clear correlation between the protein yield and change in media pH over time, where the minimal media that were the most buffered and therefore most resistant to change in pH produced the most protein. And in all three test protein cases, the difference in yield was nearly twofold between the best and worst buffering media. Thus, we propose that increasing the buffering capacity of M9 minimal media will generally lead to a similar increase for most of the proteins currently produced by this standard protein expression protocol. Moreover, we have qualitatively found that this effect also extends to deuterated M9 minimal media growths, which could lead to significant cost savings in these preparations.

Original languageEnglish (US)
Pages (from-to)11-17
Number of pages7
JournalJournal of biomolecular NMR
Issue number1-2
StatePublished - Feb 15 2019
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2019, Springer Nature B.V.


  • Buffer capacity
  • Cell growth
  • Modified M9 media
  • Protein expression


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