Enhanced methods for genetic assays in Drosophila cells

Wang, Y.; Lee, J. Y.; Housden, A.; Hottinger, E.; Housden, B. E.

doi:10.1101/2024.12.01.626214

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bioRxiv · 10.1101/2024.12.01.626214

Enhanced methods for genetic assays in Drosophila cells

Wang, Y.·Lee, J. Y.·Housden, A.·Hottinger, E.·Housden, B. E.

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Abstract

Genetic assays are an invaluable tool for both fundamental biological research and translational applications. Variable Dose Analysis (VDA) is an RNAi-based method for cell-based genetic assays that offers several advantages over approaches such as CRISPR and other RNAi-based methods including improved data quality (signal-to-noise ratio) and the ability to study essential genes at sub-lethal knockdown efficiency. Here we report the development of three new variants of the VDA method called high-throughput VDA (htVDA), VDA-plus and pooled-VDA. htVDA requires 10-fold reduced reagent volumes and takes advantage of liquid handling automation to allow higher throughput screens to be performed while maintaining high data quality. VDA-plus is a modified version of VDA that further improves data quality by 4.5-fold compared to standard VDA to allow highly sensitive detection of weak phenotypes. Finally, Pooled VDA allows greatly increased throughput by analysing multiple gene knockdowns in a single population of cells. Together, these new methods enhance the toolbox available for genetic assays, which will prove valuable in both high-and low-throughput applications. In particular, the low noise and ability of VDA to study essential genes at sub-lethal knockdown levels will support identification of novel drug-targets, among which essential genes are often enriched. While these tools have been developed in Drosophila cells, the underlying principles are transferrable to any cell culture system.

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Wang, Y., Lee, J. Y., Housden, A., Hottinger, E., Housden, B. E.. 2024-12-05. Enhanced methods for genetic assays in Drosophila cells. https://doi.org/10.1101/2024.12.01.626214

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