We used Drosophila to directly test the hypothesis that Clic genes are important for acute ethanol behavior. We confirmed two independent transposon insertions (G0472 and EY04209, Figure 2A) in the sole Clic locus in flies and subsequently backcrossed both elements into w1118 (our standard laboratory stock) to control for genetic background effects on ethanol behavior. Both transposon insertions were homozygous female and hemizygous male lethal at 25°C. Female flies heterozygous for G0472 and EY04209, however, were fertile, appeared morphologically normal and seemed generally healthy. qRT-PCR analysis showed that Clic mRNA in G0472/+ and EY04209/+ flies was reduced by approximately 40% compared to w1118 control (Figure 2B; *, one sample t tests; t(2)=27.01 and t(2)=4.95 respectively, p<0.05, n=3). G0472 and EY04209 are therefore partial loss of function mutations in Drosophila Clic.
