A general strategy for expanding polymerase function by droplet microfluidics

Andrew C. Larsen, Matthew R. Dunn, Andrew Hatch, Sujay P. Sau, Cody Youngbull, John C. Chaput

Research output: Contribution to journalArticle

44 Citations (Scopus)

Abstract

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.

Original languageEnglish (US)
Article number11235
JournalNature Communications
Volume7
DOIs
StatePublished - Apr 5 2016

Fingerprint

Microfluidics
classifying
Sorting
Polymers
Synthetic Biology
Manganese
Base Pairing
Nucleic Acids
Libraries
Copying
Optical sensors
polymers
nucleic acids
optical measuring instruments
compartments
biology
Encapsulation
manganese
templates
recovery

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Chemistry(all)
  • Physics and Astronomy(all)

Cite this

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]. https://doi.org/10.1038/ncomms11235

A general strategy for expanding polymerase function by droplet microfluidics. / Larsen, Andrew C.; Dunn, Matthew R.; Hatch, Andrew; Sau, Sujay P.; Youngbull, Cody; Chaput, John C.

In: Nature Communications, Vol. 7, 11235, 05.04.2016.

Research output: Contribution to journalArticle

Larsen, AC, Dunn, MR, Hatch, A, Sau, SP, Youngbull, C & Chaput, JC 2016, 'A general strategy for expanding polymerase function by droplet microfluidics', Nature Communications, vol. 7, 11235. https://doi.org/10.1038/ncomms11235
Larsen, Andrew C. ; Dunn, Matthew R. ; Hatch, Andrew ; Sau, Sujay P. ; Youngbull, Cody ; Chaput, John C. / A general strategy for expanding polymerase function by droplet microfluidics. In: Nature Communications. 2016 ; Vol. 7.
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