Project description:Understand the mechanisms of evolution in large-scale bio-production by tracking population dynamics leading to production decline in mevalonic acid-producing Escherichia coli. Industrial bioproduction entails growth of the production host to large bioreactors (e.g. 1-300 m3). This may put the organism at risk for generating non-producing subpopulations of genetic heterogeneity, which is not phenotypically detected at lab-scale (e.g. 2 L). To study these dynamics, we experimentally simulated these growth durations by passing mevalonic acid-producing E. coli to maintain the populations in exponential growth for 45 generations.
Project description:Long-term experiment (150 days) of Escherichia coli MC1000 with daily transfers into fresh LB medium and under three different oxygen regimes. An overnight culture of E coli (ancestor culture) was propagated in LB media with three different oxygen regimes during 150 days. One regime involved transfers under constant shaking (200rpm): Treatment A-Replica populations 1 and 2 , second regime consisted in a daily shift between shaking and static (no shaking) conditions: Treatment B-replica populations 3,4 and 5. The third regime involved transfers in constant static conditions: Treatment C-Populations 6 and 7. After 150 days and plating, 3 different colony types were selected by population (populations 6 and 7 only had 2 different morphologies). Morpholigies were marked as a, b, c in each population. cDNA was obtainned from each morphology after growing separately in LB medium, under they correspondant evolving condition until late logaritmic phase (0,6-0,7 in OD600). A total of 19 evolved forms were analyzed plus one ancestor. Each morphology was labeled in a dye-swap design and hybridized to the ancestor into genome-wide multi-strain E. coli (8x15K) microarrays.
Project description:We report identification and characterization of antibiotic persister mutants carrying characteristic mutations in the Escherichia coli rpoB gene
