We investigated a behavioral paradigm based on ethanol-induced sedation to potentially circumvent the confound of mini-w and w in eRING assays. The ethanol sedation assay we developed is based in large part on the work of others (e.g. Lasek et al., 2011; Maples and Rothenfluh, 2011; Rothenfluh et al., 2006; Schumann et al., 2011; Wen et al., 2005). In the ethanol sedation assays used in our studies, flies were placed in a 9.5 cm tall plastic food vial and trapped in the vial with a cellulose acetate Flug (Figure S3A). Ethanol solution (up to 2 ml, Figure S4A) was added to the top (exposed side) of the Flug and the vial was immediately sealed with a rubber stopper (Figure S3A). At 6-minute intervals thereafter, flies were gently tapped to the bottom of the vial and then visually scored for their ability to right themselves (i.e. stand up) in the continuous presence of ethanol vapor from the Flug. We designed our ethanol sedation assay to test multiple replicates of several genotypes in parallel (Figure S3B). Each vial of flies corresponded to
