4.7 Article

Development of a Positive Selection High Throughput Genetic Screen in Dictyostelium discoideum

期刊

出版社

FRONTIERS MEDIA SA
DOI: 10.3389/fcell.2021.725678

关键词

Dictyostelium discoideum; 5-FOA; polyglutamine; protein aggregation; genetic screen

资金

  1. National Institutes of Health [NS112191, GM119544]
  2. Diverse Scientists in Ataxia Predoctoral Fellowship from the National Ataxia Foundation

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Dictyostelium discoideum is a powerful model organism for investigating human health and disease, with high throughput genetic screens. However, limitations such as time-consuming screening processes have restricted genomic coverage. By coupling chemical mutagenesis with the 5-fluoroorotic acid counterselection system, mutations in genes of interest can be enriched efficiently, offering potential for broader applications across different biological systems.
The cellular slime mold Dictyostelium discoideum is a powerful model organism that can be utilized to investigate human health and disease. One particular strength of Dictyostelium is that it can be utilized for high throughput genetic screens. For many phenotypes, one limitation of utilizing Dictyostelium is that screening can be an arduous and time-consuming process, limiting the genomic depth one can cover. Previously, we utilized a restriction enzyme-mediated integration screen to identify suppressors of polyglutamine aggregation in Dictyostelium. However, due to the time required to perform the screen, we only obtained & SIM;4% genome coverage. Here we have developed an efficient screening pipeline that couples chemical mutagenesis with the 5-fluoroorotic acid counterselection system to enrich for mutations in genes of interest. Here we describe this new screening methodology and highlight how it can be utilized for other biological systems.

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