Development of a novel continuous culture device for experimental evolution of bacterial populations
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The availability of a robust and reliable continuous culture apparatus that eliminates wall growth problems would lead to many applications in the microbial field, including allowing genetically engineered strains to recover high fitness, improving biodegradation strains, and predicting likely antibiotic resistance mechanisms. We describe the design and implementation of a novel automated continuous culture machine that can be used both in time-dependent mode (similar to a chemostat) and turbidostat modes, in which wall growth is circumvented through the use of a long, variably divisible tube of growth medium. This tube can be restricted with clamps to create a mobile growth chamber region in which static portions of the tube and the associated medium are replaced together at equal rates. To functionally test the device as a tool for re-adaptation of engineered strains, we evolved a strain carrying a highly deleterious deletion of Elongation Factor P, a gene involved in translation. In 200 generations over 2 weeks of dilution cycles, the evolved strain improved in generation time by a factor of three, with no contaminations and easy manipulation.
KeywordsExperimental evolution Continuous culture Turbidostat Natural selection Adaptation Metabolic engineering Biodegradation
We thank Phillippe Marlière for inspiration, Paul Schimmel for encouragements in the initial stages of the project, and Daniel Dykhuizen for critical reading of the manuscript. V d C-L work was funded in part by grant MCB-0128901 from the National Science Foundation.
- de Crécy E (2005) Continuous culture apparatus with mobile vessel allowing selection of fitter cell variants. WO/2005/083052Google Scholar
- Dykhuizen DE, Hartl DL (1983) Selection in chemostats. Microbiol Rev 47:150–168Google Scholar
- Helling RB, Kinney T, Adams J (1981) The maintenance of plasmid-containing organisms in populations of Escherichia coli. J Gen Microbiol 123:129–141Google Scholar
- Larsen DH, Dimmick RL (1964) Attachment and growth of bacteria on surfaces of continuous culture vessels. J Bacteriol 88:1380–1387Google Scholar
- Monod J (1950) La technique de culture continue. Théorie et applications. Ann Inst Pasteur 19:390–410Google Scholar
- Mutzel R, Mazel D, Marlière P (2003) Method for obtaining cells with new properties WO 03/004656Google Scholar
- Vidal O, Longin R, Prigent-Combaret C, Dorel C, Hooreman M, Lejeune P (1998) Isolation of an Escherichia coli K-12 mutant strain able to form biofilms on inert surfaces: involvement of a new ompR allele that increases curli expression. J Bacteriol 180:2442–2449Google Scholar
- Wahl LM, Gerrish PJ, Saika-Voivod I (2002) Evaluating the impact of population bottlenecks in experimental evolution. Genetics 162:961Google Scholar