Generation of Or7a Knockout Mutants

Accelerated homologous recombination was performed according to Baena-Lopez LA et al. (Baena-Lopez et al., 2013 (link)). Briefly, P-element insertion lines containing the Or7a knockout construct were crossed to hs-Flp, hs-SceI (BS#25679) and heat-shocked at 48 and 72 hr after egg-laying (1 hr duration each time). Female progeny with mottled eyes were crossed to ubi-Gal4[pax-GFP] (Baena-Lopez et al., 2013 (link)) in order to select against flies containing non-homologous recombination events. Stocks were generated from candidate flies that contained both w⁺ and GFP markers. Or7a mutants were verified by single sensillum recordings and PCR (Figure 4F, G, Figure 4—figure supplement 3). In order to identify the ab4 sensillum, 30 μl of geosmin (Sigma #16423-19-1), an odor that specifically activates only ab4B (Or56a) (Stensmyr et al., 2012 (link)), was used (Figure 4F,G).
Primers used for verification: G4polyA_FOR: TCG ATA CCG TCG ACT AAA GCC; gOr7a_REV:TCG CCG TTG AGT TTT CAG AG
The Or7a-Gal4 knockin was generated by co-injection of the pRIV-Gal4 donor plasmid (Baena-Lopez et al., 2013 (link)) with PhiC31 integrase to target GAL4 to the attP site within the knockout locus, as described in (Baena-Lopez et al., 2013 (link)).