Here, we describe a fast and cost-effective procedure to generate a large array of mutant proteins and immediately screen for those with altered protein function. This protocol is a modification from three existing approaches: fusion PCR, Saccharomyces cerevisiae in-yeast recombination, and semi-quantitative growth assays. We also describe a mating step to reduce the occurrence of false positive findings due to ectopic mutations. The only requirement is that the protein elicits a phenotype in Saccharomyces cerevisiae.
Keywords: Biotechnology and bioengineering; Cell-based Assays; Model Organisms; Molecular Biology.
Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.