|DRSC: JAK-STAT Pathway and dsRNA Consistency - BioAssay Summary
Abstract: Background: High-throughput screening using RNAi is a powerful gene discovery method but is often complicated by false positive and false negative results. Whereas false positive results associated with RNAi reagents has been a matter of extensive study, the issue of false negatives has received less attention. ..more
Title: False negative rates in Drosophila cell-based RNAi screens
Abstract: Background: High-throughput screening using RNAi is a powerful gene discovery method but is often complicated by false positive and false negative results. Whereas false positive results associated with RNAi reagents has been a matter of extensive study, the issue of false negatives has received less attention.
Results: We performed a meta-analysis of several genome-wide, cell-based Drosophila RNAi screens, together with a more focused RNAi screen, and conclude that the rate of false negative results is at least 8%. Further, we demonstrate how knowledge of the cell transcriptome can be used to resolve ambiguous results and how the number of false negative results can be reduced by using multiple, independently-tested RNAi reagents per gene.
Conclusions: RNAi reagents that target the same gene do not always yield consistent results due to false positives and weak or ineffective reagents. False positive results can be partially minimized by filtering with transcriptome data. RNAi libraries with multiple reagents per gene also reduce false positive and false negative outcomes when inconsistent results are disambiguated carefully.
We selected dsRNAs with Z-scores with an absolute value of 2 or greater across both replicates, which in this case included dsRNAs targeting 24 genes. We then compared these to the Z-scores of the other dsRNAs in the K/P set that target the same gene and transcripts. In some cases, the scores obtained with all dsRNAs directed against a particular gene were consistent, whereas in other cases, some dsRNAs directed against a single gene were phenotypically inconsistent. We categorized the results of dsRNAs into three categories: In category 1, all dsRNAs directed against a given gene were hits. In category 2, at least 2 dsRNAs were hits but there was at least one which did not score significantly. In category 3, only 1 dsRNA directed against a gene yielded a significant result. Out of 24 genes, 5 had positive results for all dsRNAs (category 1), 4 were in category 2, and 15 were in category 3.
Target Type: ORGANISM
Pref Name: Drosophila melanogaster
Description: Fruit Fly
Organism: Drosophila melanogaster
Tax ID: 7227
Confidence: Target assigned is non-molecular
Relationship Type: Non-molecular target assigned
Data Table (Concise)