High-Throughput Image-Based Screening of Pooled Genetic Variant Libraries
Emanuel, George Alexander
MetadataShow full item record
AbstractImage-based, high-throughput screening of pooled libraries of genetic perturbations will greatly advance our understanding of biological systems and facilitate many biotechnology applications. Here we report a high-throughput screening method that allows diverse genotypes and the corresponding phenotypes to be imaged in numerous individual cells. To facilitate genotyping by imaging, we introduced barcoded genetic variants into cells, each cell carrying a single genetic variant connected to a nucleic-acid barcode. We then performed live-cell imaging to determine the phenotype of each cell and massively multiplexed FISH imaging to measure the barcode expressed in the same cell for genotype identification. We demonstrated the utility of this method by screening genetic variants of the fluorescent protein YFAST. We imaged 20 million cells expressing ~60,000 YFAST mutants and identified novel YFAST variants that are both brighter and more photostable than the original protein.
Citable link to this pagehttp://nrs.harvard.edu/urn-3:HUL.InstRepos:40050109
- FAS Theses and Dissertations