Successful in vivo directed evolution by PREDCEL and PACE requires the thorough consideration of experimental parameters, e.g. phage propagation times, culture dilution rates and inducer/inhibitor concentrations. We employed extensive ODE-based and stochastic modeling to identify the most sensitive parameters and adapt our experiments accordingly. First, we calibrated our models using phage propagation experiments from our wet lab complemented with literature data. Simulations showed that the phage titer is highly sensitive to culture dilution rates. We simulated batch times and transfer volumes for PREDCEL and corresponding flow rates for PACE to determine optimized conditions for gene pool selection while avoiding phage washout. We also estimated phage titer monitoring intervals for cost and labor efficient QC/monitoring as well as inducer/inhibitor concentrations required to express the required mutagenic polymerases. Finally, we provide a web-based, fully interactive modeling platform that not only informed our wet lab experiments, but enables future iGEM teams to efficiently build on our work.
Simulations of phage and E. coli titer support both PREDCEL and PACE by helping to choose a set of experimental parameters that is both efficient in terms of directed evolution and in terms of usability.