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Validating RNAi Phenotypes in Drosophila Using a Synthetic RNAi-Resistant Transgene

Abstract : RNA interference (RNAi) is a powerful and widely used approach to investigate gene function, but a major limitation of the approach is the high incidence of non-specific phenotypes that arise due to off-target effects. We previously showed that RNAi-mediated knock-down of pico, which encodes the only member of the MRL family of adapter proteins in Drosophila, resulted in reduction in cell number and size leading to reduced tissue growth. In contrast, a recent study reported that pico knockdown leads to tissue dysmorphology, pointing to an indirect role for pico in the control of wing size. To understand the cause of this disparity we have utilised a synthetic RNAi-resistant transgene, which bears minimal sequence homology to the predicted dsRNA but encodes wild type Pico protein, to reanalyse the RNAi lines used in the two studies. We find that the RNAi lines from different sources exhibit different effects, with one set of lines uniquely resulting in a tissue dysmorphology phenotype when expressed in the developing wing. Importantly, the loss of tissue morphology fails to be complemented by co-overexpression of RNAi-resistant pico suggesting that this phenotype is the result of an off-target effect. This highlights the importance of careful validation of RNAi-induced phenotypes, and shows the potential of synthetic transgenes for their experimental validation.
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https://hal.inrae.fr/hal-03071690
Contributor : Vincent Jonchere <>
Submitted on : Wednesday, December 16, 2020 - 10:17:06 AM
Last modification on : Tuesday, April 20, 2021 - 9:37:15 AM

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Vincent Jonchère, Daimark Bennett. Validating RNAi Phenotypes in Drosophila Using a Synthetic RNAi-Resistant Transgene. PLoS ONE, Public Library of Science, 2013, 8 (8), pp.e70489. ⟨10.1371/journal.pone.0070489⟩. ⟨hal-03071690⟩

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