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A genetic multicolor cell-labeling technique for Droshophila melanogaster, Flybow, is described and applied to the study of neural circuits. This method implements a variant of the mouse Brainbow strategy in combination with specific neuronal targeting using the Gal-4–upstream activating sequence system to select for membrane-tethered fluorescent proteins. Also in this issue, Hampel et al. report a similar strategy, Drosophila Brainbow, to select for epitope-tagged proteins detectable via immunofluorescence.
A genetic multicolor cell-labeling technique for Droshophila melanogaster, Drosophila Brainbow, is described and applied to the study of neural circuits. This method implements a variant of the mouse Brainbow strategy in combination with specific neuronal targeting using the Gal-4–upstream activating sequence system to select for epitope-tagged proteins detectable with immunofluorescence. Also in this issue, Hadjieconomou et al. develop a similar strategy, Flybow, to select for membrane-tethered fluorescent proteins.
Changing the codon sequence in Caenorhabditis elegans genes allows fine-tuning of transgene expression from high to low expression. The same strategy is likely applicable for Drosophila melanogaster and Saccharomyces cerevisiae.
By methylating the phosphate groups of PtdIns(3,4,5)P3 researchers can load this lipid more efficiently into a mass spectrometer and thus this lipid can be quantified in the presence of an internal synthetic standard.
A genetic platform allows refinement of tissue-specific expression using the upstream activating sequence–GAL4 system in Drosophila melanogaster, facilitating the segmentation of complex expression patterns and allowing GAL4 expression patterns to be repurposed.
Microscope control software allowed automatic machine learning-based detection of rare events in living cells and unattended operation of complex imaging assays. Performance was demonstrated by detailed analysis of processes during transient mitotic stages.
Stimulated Raman scattering (SRS) microscopy is a quantitative, label-free imaging method to map fat distribution and accumulation with high spatial resolution and sensitivity at both cellular and organism levels.
An optogenetic illumination system based on the use of a digital micromirror device and video tracking software is reported, which allows real-time light delivery with high spatial resolution to specified targets in freely moving Caenorhabditis elegans. Also in this issue, Stirman et al. report a similar illumination system using a liquid crystal display projector. Both methods allow optogenetic perturbation of a variety of neural circuits in the behaving worm.
An optogenetic illumination system based on the use of a liquid crystal display projector and video tracking software is reported, which allows real-time multispectral light delivery with high spatial resolution to specified targets in freely moving Caenorhabditis elegans. Also in this issue, Leifer et al. report a similar illumination system using a digital micromirror device. Both methods allow optogenetic perturbation of a variety of neural circuits in the behaving worm.
In vivo calcium imaging at multiple depths simultaneously is shown using multifocal two-photon microscopy and spatiotemporal multiplexing. This technique involves scanning the sample with multiple beams in parallel at different axial planes and is applied to monitor neuronal network activity in multiple cortical layers of an anesthetized mouse.
Single-molecule fluorescence experiments with microsecond time resolution are made possible using a photoprotection cocktail that reduces dye blinking and bleaching with a combination of dissolved oxygen, a triplet quencher and a free-radical scavenger.
By individually replacing 16 yeast genes encoding ABC transporters by GFP, mating and selecting for strains with accumulated mutations the authors create a Green Monster, a strain with deletions in all 16 genes.
