biostudies-literatureStankovic DDeutsche ForschungsgemeinschaftNIH HHSe202201801https://www.ebi.ac.uk/biostudies/studies/S-EPMC9711858biostudies-literatureUnknown6(2)Post-transcriptional gene silencing using double-stranded RNA has revolutionized the field of functional genetics, allowing fast and easy disruption of gene function in various organisms. In <i>Drosophila</i>, many transgenic RNAi lines have been generated in large-scale efforts, including the <i>Drosophila</i> Transgenic RNAi Project (TRiP), to facilitate in vivo knockdown of virtually any <i>Drosophila</i> gene with spatial and temporal resolution. The available transgenic RNAi lines represent a fundamental resource for the fly community, providing an unprecedented opportunity to address a vast range of biological questions relevant to basic and biomedical research fields. However, caution should be applied regarding the efficiency and specificity of the RNAi approach. Here, we demonstraLife science alliance<i>Drosophila</i> pVALIUM10 TRiP RNAi lines cause undesired silencing of Gateway-based transgenes.PMC9711858Germany&apos;s Excellence Strategy – CECAD, EXC 2030 – 390661388P40 OD010949Csordas GStankovic DUhlirova Mfalse<i>Drosophila</i> pVALIUM10 TRiP RNAi lines cause undesired silencing of Gateway-based transgenes.Post-transcriptional gene silencing using double-stranded RNA has revolutionized the field of functional genetics, allowing fast and easy disruption of gene function in various organisms. In <i>Drosophila</i>, many transgenic RNAi lines have been generated in large-scale efforts, including the <i>Drosophila</i> Transgenic RNAi Project (TRiP), to facilitate in vivo knockdown of virtually any <i>Drosophila</i> gene with spatial and temporal resolution. The available transgenic RNAi lines represent a fundamental resource for the fly community, providing an unprecedented opportunity to address a vast range of biological questions relevant to basic and biomedical research fields. However, caution should be applied regarding the efficiency and specificity of the RNAi approach. Here, we demonstra2023-01-01T00:00:00Z2023 Feb2026-05-29T04:34:04.946Z2025-04-04T19:37:59.641ZS-EPMC97118583644652210.26508/lsa.202201801