A Normalization Protocol Reduces Edge Effect in High-Throughput Analyses of Hydroxyurea Hypersensitivity in Fission...

Edge effect denotes better growth of microbial organisms situated at the edge of the solid agar media. Although the precise reason underlying edge effect is unresolved, it is generally attributed to greater nutrient availability with less competing neighbors at the edge. Nonetheless, edge effect constitutes an unavoidable confounding factor that results in misinterpretation of cell fitness, especially in high-throughput screening experiments widely employed for genome-wide investigation using microbial gene knockout or mutant libraries. Here, we visualize edge effect in high-throughput high-density pinning arrays and report a normalization approach based on colony growth rate to quantify drug (hydroxyurea)-hypersensitivity in fission yeast strains. This normalization procedure improved the accuracy of fitness measurement by compensating cell growth rate discrepancy at different locations on the plate and reducing false-positive and -negative frequencies. Our work thus provides a simple and coding-free solution for a struggling problem in robotics-based high-throughput screening experiments.