Genetic Interaction Mapping in <i>Schizosaccharomyces pombe</i> Using the Pombe Epistasis Mapper (PEM) System and a ROTOR HDA Colony Replicating Robot in a 1536 Array Format

摘要

This protocol describes an optimized high-throughput procedure for generating double deletion mutants in Schizosaccharomyces pombe using the colony replicating robot ROTOR HDA and the PEM (pombe epistasis mapper) system. The method is based on generating high-density colony arrays (1536 colonies per agar plate) and passaging them through a series of antidiploid and mating-type selection (ADS-MTS) and double-mutant selection (DMS) steps. Detailed program parameters for each individual replication step are provided. Using this procedure, batches of 25 or more screens can be routinely performed.