A high-throughput differential chemical genetic screen uncovers genotype-specific compounds altering plant growth

The identification of chemical compounds regulating plant growth in a genetic context can greatly enhance our understanding of biological mechanisms. Here, we have developed a high-throughput phenotype-directed chemical screening method in plants to compare two genotypes and identify small molecules inducing genotype-specific phenotypes. We used <i>Arabidopsis thaliana</i> wild type and <i>mus81</i>, a DNA repair mutant, and screened off-patent drugs from the Prestwick library to selectively identify molecules affecting <i>mus81</i> growth. We developed two complementary convolutional neural networks (CNN)-based image segmentation and classification programs to quantify <i>Arabidopsis</i> seedling growth. Using these approaches, we detected that about 10% of Prestwick molecules cause altered growth in both genotypes, suggesting their toxic effects on plant growth. We identified three Prestwick molecules specifically affecting <i>mus81</i>. Overall, we developed a straightforward, accurate, and adaptable methodology for performing high-throughput screening of chemical libraries in a time-efficient manner, accelerating the discovery of genotype-specific chemical regulators of plant growth.