Full metadata
Polymerases that synthesize artificial genetic polymers hold great promise for advancing future applications in synthetic biology. However, engineering natural polymerases to replicate unnatural genetic polymers is a challenging problem. Here we present droplet-based optical polymerase sorting (DrOPS) as a general strategy for expanding polymerase function that employs an optical sensor to monitor polymerase activity inside the microenvironment of a uniform synthetic compartment generated by microfluidics. We validated this approach by performing a complete cycle of encapsulation, sorting and recovery on a doped library and observed an enrichment of ∼1,200-fold for a model engineered polymerase. We then applied our method to evolve a manganese-independent α-L-threofuranosyl nucleic acid (TNA) polymerase that functions with >99% template-copying fidelity. Based on our findings, we suggest that DrOPS is a versatile tool that could be used to evolve any polymerase function, where optical detection can be achieved by Watson-Crick base pairing.
- Larsen, Andrew (Author)
- Dunn, Matthew (Author)
- Hatch, Andrew (Author)
- Sau, Sujay (Author)
- Youngbull, Cody (Author)
- Chaput, John (Author)
- Biodesign Institute (Contributor)
Larsen, A. C., Dunn, M. R., Hatch, A., Sau, S. P., Youngbull, C., & Chaput, J. C. (2016). A general strategy for expanding polymerase function by droplet microfluidics. Nature Communications, 7, 11235. doi:10.1038/ncomms11235
- 2017-06-01 04:22:15
- 2021-11-03 12:18:27
- 3 years ago