User-defined single pot mutagenesis using unamplified oligo pools
Abstract
User-defined mutagenic libraries are fundamental for applied protein engineering workflows. Here we show that unamplified oligo pools can be used to prepare site saturation mutagenesis libraries from plasmid DNA with near-complete coverage of desired mutations and few off-target mutations. We find that oligo pools yield higher quality libraries when compared to individually synthesized degenerate oligos. We also show that multiple libraries can be multiplexed into a single oligo pool, making preparation of multiple libraries less expensive and more convenient. We provide software for automatic oligo pool design that can generate mutagenic oligos for saturating or focused libraries.
Document Details
- Document Type
- Pub Defense Publication
- Publication Date
- Jan 01, 2019
- Source ID
- 10.1093/protein/gzz013
Entities
People
- Alexandra N. Borelli
- Angélica V. Medina-cucurella
- Jesús Beltrán
- Matthew S Faber
- Monica B Kirby
- Paul J Steiner
- Sean Cutler
- Timothy A Whitehead
Organizations
- Defense Advanced Research Projects Agency
- Michigan State University
- National Institutes of Health
- United States Department of Defense
- University of California
- University of Colorado