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. 2021 Oct 20;15:732183. doi: 10.3389/fncir.2021.732183

FIGURE 1.

FIGURE 1

Bitbow1 design and characterization of labeling properties. (A) Schematic of Bitbow1 design. Five spectrally distinct FPs are separated by five pairs of reversely positioned orthogonal FRT sites. The mTFP/FRT-F14 module is shown in the dashed box. The FP’s open reading frame (ORF) is positioned in the reverse direction, corresponding to a default OFF state (‘0’). Upon Flp induced recombination, the FP’s ORF may spin to the forward direction for Gal4 driven expression, corresponding to an ON state (‘1’). (B) 31 Bitbow color codes could be generated in a single Bitbow1 brain. (C) A hsFlp;elav-Gal4 driver fly was crossed to the mBitbow1.0 fly to examine the offspring expression in the nervous system upon heat-shock induced Flp activity. Experimental setups of adult heat shock-induced labeling. (D) Maximum intensity projection overview of an adult heat-chocked brain. (E) Left panel, enlarged boxed region in panel (D) showed individual neurons are labeled in distinct colors, i.e., Bitbow codes. Right panel, Bitbow codes of four selected optic lobe neurons’ somas or terminals. (F) Experimental setups of generating heat shock-induced Bitbow labeling in 3rd instar brains. (G) Maximum intensity projection overview of a 3rd instar larvae heat-chocked brain. Inset, the enlarged boxed region showed clusters of cells labeled in the same colors. Asterisk indicates a neuroblast. (H) Quantification of occurrence frequencies of each Bitbow color. Among all quantified clusters, the fraction of clusters containing each Bitbow color were displayed. 787 clusters from 6 brains are included. Each dot on the graph represents quantification from one brain. Scale bars: (D,G) 50 μm, (E) 10 μm.