Uas mcd8 chrfp

Manufactured by Bloomington Drosophila Stock Center

Sourced in United States

The UAS-mCD8-ChRFP is a genetic construct that expresses a membrane-targeted, red fluorescent protein (ChRFP) under the control of the Upstream Activating Sequence (UAS) promoter. This construct is commonly used for cell labeling and imaging in Drosophila research.

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5 protocols using uas mcd8 chrfp

Genetic Tools for Drosophila Research

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QUAS-DtA^ts,¹² (link)WP-QF2,¹² (link)C10-Gal4 (gifted by E. Hafen), UAS-lacZ-RNAi,⁵⁵ (link) and UAS-lacZ (gifted by C. Goodman) have been characterized in previous studies. TkR86C-T2A-Gal4 was gifted by S. Kondo.²³ (link)
TkR86C-T2A-lexA was a gift from Rao.²⁴ (link)
UAS-rpr (#5824), UAS-GFP (#1521), rn-Gal4 (#51280), nSyb-Gal4 (#51635), QUAS-mCD8-GFP (#30002, #30003), UAS-nls-mCherry (#38425), UAS-mCD8-ChRFP (#27391), UAS-mCD8-ChRFP (#27392), LexAop-nls-GFP (#29954), LexAop-mCD8-GFP (#32203), UAS-Tk-RNAi (#25800), and Tk-T2A-Gal4 (#84693) were obtained from the Bloomington Drosophila Stock Center. UAS-TkR86C-RNAi 1 (v107090), UAS-TkR86C-RNAi 2 (v13392), and RNAi lines used for GPCR screening (Table S1) were obtained from the Vienna Drosophila Resource Center.

Kashio S., Masuda S, & Miura M. (2023). Involvement of neuronal tachykinin-like receptor at 86C in Drosophila disc repair via regulation of kynurenine metabolism. iScience, 26(9), 107553.

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Drosophila Neurogenetic Toolkit Protocols

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Flies were kept on a standard corn meal, corn syrup, yeast, and agar medium at room temperature. UAS-mCD8-ChRFP (#27392), vGlut-Gal4 (#26160), and Gad1-Gal4 (#47140) were newly obtained from the Bloomington Drosophila Stock Center (Indiana, USA) for these experiments. The UAS-Ca-alpha1T-IR line (#48008) was obtained from the Vienna Drosophila RNAi Center. EP line G1047 was obtained from Genexel. c465-Gal4 and 210y-Gal4 were gifts from J. Douglas Armstrong28 (link). The following stocks were all described previously: elav-Gal429 (link), elav-GS-Gal419 (link), Cha-Gal430 (link), 104y-Gal431 (link), c309-Gal432 (link), MB247-Gal433 (link), pdf-Gal434 (link), TH-Gal435 (link), GMR-Gal436 (link), clk8.0-Gal437 (link), dilp2-Gal438 (link), Tdc2-Gal4 and TRH-Gal439 (link), c161-Gal4 and c232-Gal440 (link), and c929-Gal4 and 386Y-Gal441 (link).

Jeong K., Lee S., Seo H., Oh Y., Jang D., Choe J., Kim D., Lee J.H, & Jones W.D. (2015). Ca-α1T, a fly T-type Ca2+ channel, negatively modulates sleep. Scientific Reports, 5, 17893.

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Drosophila Nerve Injury and Transgene Expression

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Flies were grown on standard cornmeal, molasses food at 25°C. Males were typically used. Strains included appl-GAL4, Hemese-GAL4, repo-GAL4, UAS-GAL4, UAS-mCD8-GFP, UAS-mCD8-ChRFP, and LexAop-mCD8-GFP, UAS-bskDN, UAS-hepCA, and UAS-Lifeact-Ruby27 (link)28 (link) (Bloomington Drosophila Stock Center (BDSC), Indiana University (Bloomington, IN)). Other lines were UAS-EB1-RFP (Dr. M. Rolls, U. Michigan, Ann Arbor, MI), LexA-CD4-GFP and LexA/nSynaptobrevin46 (link), elav-GS32 (link), tubulin-GS (Dr. Scott Pletcher, U Michigan, Ann Arbor, MI). Animals were collected over a 2h period then aged 5h for 6h, 24h for 1d, and 72h for 3d animals. Flies on RU486 (Mifepristone, Sigma Aldrich, cat# M8046) were maintained on food supplemented with drug32 (link). Briefly, for each food vial, 50 ul of RU486 stock solution [4.0 mg/ml in 100% ethanol] was pipetted onto the medium and allowed to penetrate the food overnight. UAS-transgenes showed robust expression throughout the wing within 18h of being placed on RU486-containing food, and visible transgene expression (hinge region close to the body of animal) by 6h32 (link)47 (link). Bottles of animals were cleared, collected over a 2h period and put on RU486 or vehicle food and aged for nerve injury.

Soares L., Parisi M, & Bonini N.M. (2014). Axon Injury and Regeneration in the Adult Drosophila. Scientific Reports, 4, 6199.

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Live Imaging of Drosophila Morphogenesis

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Flies were raised at room temperature on standard fly food. The following fly stocks were obtained from the Bloomington Drosophila Stock Center: {en2.4-Gal4}e16E (BL30564); w; P{UAS-mCD8.ChRFP} (BL 27392). w; btl-Gal4 UAS-CAAX-mCherry/TM6b was obtained from S. Hayashi (RIKEN Center for Developmental Biology, Kobe, Japan). Genotypes used for live imaging were ;en-Gal4/CyO; UAS-cd8mCherry for leading edge cells during dorsal closure and btl-Gal4 UAS-Cherry-CAAX/TM6b for terminal tracheal cells. After egg-laying overnight at 25°C, embryos were dechorionated in thin bleach for 1 min and mounted on a coverslip with heptane glue and covered with water. Embryos at the end of stage 14 and 16 were selected for imaging dorsal closure and tracheal cells, respectively. Live imaging was performed at room temperature on an inverted TCS-sp5 confocal microscope (Leica) equipped with a 63× 1.4 NA plan Apo oil immersion objective and Hybrid detectors HyD, using Leica software. Stacks of 7–10 z sections (0.5 or 0.7 µm Z step size) were taken every 15 s for 10 min. Using ImageJ, a maximum intensity z-projection was then applied to use the plugin Filopodyan for filopodia reconstruction.

Urbančič V., Butler R., Richier B., Peter M., Mason J., Livesey F.J., Holt C.E, & Gallop J.L. (2017). Filopodyan: An open-source pipeline for the analysis of filopodia. The Journal of Cell Biology, 216(10), 3405-3422.

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Overexpression studies of Notch signaling components

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Overexpression studies were performed using the Gal4/UAS system^{36 (link)}. We used the following UAS lines: UAS-Dl, UAS-Ser, UAS-mCD8-ChRFP (Bloomington Drosophila Stock Center; BDSC), UAS-fwdIR, UAS-sktlIR, UAS-plc-21IR, UAS-dgkIR, UAS-gfpIR (Vienna Drosophila RNAi Center). UAS-pld38, and UAS-rdgA/dgk were a generous gift from Padinjat Raghu (National Centre for Biological Sciences, Tata Institute of Fundamental Research, India). We also used the following Gal4 lines: pnr-Gal4, en-Gal4 and sca-Gal4 (BDSC). We also used the following strains Dl^[Rev10], Ser^[RX82], Spdo^G104, Ap-2α^[40–31], numb¹⁵^{37 (link)}. SpdoCh2GFP3, neur-nlsFP670, NiGFP were a generous gift from François Schweisguth (Institut Pasteur, France). UAS-lipinwt was a generous gift from Michael Lehmann (University of Arkansas, USA). All phenotypes were analysed at 25 °C unless stated otherwise.

Medina-Yáñez I., Olivares G.H., Vega-Macaya F., Mlodzik M, & Olguín P. (2020). Phosphatidic acid increases Notch signalling by affecting Sanpodo trafficking during Drosophila sensory organ development. Scientific Reports, 10, 21731.

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