Phage-Assisted Continuous Evolution (PACE): A Guide Focused on Evolving Protein–DNA Interactions
Serban C. Popa, Ichiro Inamoto, B W. Thuronyi, Jumi A. Shin
Abstract
evolution, where the protein under selective pressure undergoes continuous mutagenesis with little-to-no input from the researcher once the system is started. However, continuous evolution methods can be challenging to implement and a daunting investment of time and resources. Our intent is to provide basic information and helpful suggestions that we have gained from our experience with bacterial phage-assisted continuous evolution (PACE) toward the evolution of proteins that bind to a specific DNA target. We discuss factors to consider before adopting PACE for a given evolution scheme with focus on the PACE bacterial one-hybrid selection system and what optimization of a PACE selection circuit may look like using the evolution of the DNA-binding protein ME47 as a case study. We outline different types of selection circuits and techniques that may be added onto a basic PACE setup. With this information, researchers will be better equipped to determine whether PACE is a valid strategy to adopt for their research program and how to set up a valid selection circuit.