Local interneurons (LNs) of the glutamatergic ventrolateral cluster derive from a novel lineage. (A) Dual binary expression system to label (A1) LNs using Gal4-OK371 > UAS-mCD8::GFP and (A2) projection neurons (PNs) using GH146-LexA::GAD > rCD2::GFP. (A3, A4) A clear separation of the LNs (green) and PNs (magenta) can be seen. (B1-B4) tub-LexA::GAD/+ (or Y); FRTG13, Gal4-GH146, UAS-mCD8/FRT G13, hsFLP, tub-GAL80 ; lexAop-rCD2::GFP/+. Dual MARCM clone induced at 0-4 ALH. Clone labelled with (B1) Tub is (demarcated with yellow dots) and (B2) Gal4-GH146 (yellow dots) is labelled with (B3) anti-GABA; (B4) Merged image of the different channels in B1-B3. (C1-C4) tub-LexA::GAD/+ or Y; FRTG13, Gal4-GH146, UAS-mCD8/FRT G13, hsFLP, tub-GAL80 ; lexAop-rCD2::GFP/+. Dual MARCM clone induced at 0-4 ALH. The clone labelled with (C1) tub (yellow dots) is not labelled with (C2) Gal4-GH146. (C1, C4) Primary neurite bundle from the cell bodies (magenta arrow). (C3) The clonal cells are not labelled with anti-GABA. (C1, C4) Arbours of these cells cross to the contralateral lobe via the commissure (yellow arrow). (D1-D3) tub-LexA::GAD/+ or Y; FRT G13, hsFLP, tub-GAL80/UASmCD8::GFP, Gal4-OK371, FRTG13; lexAop-rCD2::GFP/+. Dual MARCM clone generated by heat shock at 24 ALH. The clone labelled with (D1, D3) tub completely overlaps that labelled with (D2, D3) Gal4-OK371. Glutamatergic ventrolateral cluster (cyan dotted line). (A4) Antennal lobes (demarcated by white dots). D = dorsal; L = lateral. Scale bars = 10 μm.