Specific mutations are generated in Drosophila homologs of human genes to study phenotypic consequences in vivo. Gene function can be reduced or silenced using CRISPR-Cas9 and RNAi interference, or overexpressed via transgenes. In other words, reverse genetics first observes a gene, then analyzes a phenotype.

Reverse Genetics

In forward genetics, mutations are generated at random, traditionally with ethyl methanesulfonate (EMS). The mutant progeny are then screened for any aberrant phenotype or behavior. Finally, the DNA of mutants displaying a similar phenotype or behavior are sequenced and compared for common polymorphisms. In other words, forward genetics first observes a phenotype, then analyzes a gene.

Forward Genetics

One of the strategies used to identify human disease-causing genes is whole-genome/exome sequencing of patients and their families. However, this strategy has limited statistical power. Through the diagnostic strategy, Drosophila homologs of human disease-causing genes are identified and knocked out via the GAL4-UAS system. If the knockout fly displays a change in phenotype, and expressing wild-type UAS-human-cDNA rescues the aberrant phenotype, then there is evidence that the selected gene causes the associated disease.

Diagnostic Strategy

- Reverse genetics
- Forward genetics
- Diagnostic strategy

Pomona College

 Drosophila melanogaster (fruit flies) are valuable model organisms for studying mechanisms underlying development and human disease. Like C. elegans, Drosophila are favored models due to their short generation time, high rate of reproduction, and convenient maintenance within the laboratory. Around 65% of genes that cause disease in humans have functional homologs in flies, and 38 of 59 human neurological genes have orthologs in flies. Additionally, the Drosophila genome has been fully sequenced, allowing for more precision when manipulating genes.

Drosophila melanogaster (fruit fly)

Ugur, B., Chen, K., & Bellen, H. J. (2016). Drosophila tools and assays for the study of human diseases. Disease Models & Mechanisms, 9(3), 235–244. https://doi.org/10.1242/dmm.023762

Drosophila tools and assays for the study of human diseases (2016)

Genetic tools for Drosophila

 The chemotaxis assay is used to study C. elegans olfactory behavior. The odorant or chemical of study is applied to one side of the agar plate, with a control chemical applied to the opposite side. Sodium azide is added to both sides to immobilize the worms upon reaching their chosen odorant. Worms are placed equidistant from either chemical and allowed to chemotax, or roam freely, for a set period of time (roughly an hour to ensure the sodium azide immobilizes all worms). A chemotaxis index is calculated by subtracting the number of worms on the control zone from the number of worms on the odorant zone, divided by the total number of worms. Therefore, a chemotaxis close to 1 indicates a strong preference for the odorant, while a value close to -1 indicates either a strong repulsion to the odorant, or a lack of preference in general. This assay is also applicable to Drosophila larvae, and can be adapted to study more complex behaviors such as associative learning.

Chemotaxis Assay

- Electroretinogram (ERG)
- Neuromuscular junction electrophysiology
- Giant-fiber-system recordings
- Wing injury assay


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