Uas gfp nls

Manufactured by Bloomington Drosophila Stock Center

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The UAS-GFP.nls is a genetic construct that contains the Green Fluorescent Protein (GFP) gene with a nuclear localization signal (nls). This construct is designed to be used in conjunction with the GAL4/UAS system for targeted gene expression in Drosophila melanogaster.

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UAS-GFP (63 citations) UAS-NLS-NESP12-GFP (3 citations)

20 protocols using uas gfp nls

Genetic Toolkit for Sensory Neuron Analysis

Cited 1 time

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Bx^MS1096-GAL4, LzGal4,UAS-mCD8::GFP.L, UAS-Ras^V12, spag^k12101/CyO,UAS-GFP.nls, santa-maria¹, and the deficiency lines Df(2L)Exel7031/CyO and Df(2L)ED479, P(3′.RS5+3.3′) ED479/SM6a, which cover the santa-maria locus, were obtained from the Bloomington Stock Center (USA). UAS-grim was obtained from the Vanderbilt University Medical Center, Nashville, Tennessee 37232). The vkg^G00454-GFP-trap line was obtained from from FlyTrap (Quiñones-Coello et al., 2007 (link)). The hs-santa-maria strain was a kind gift of Craig Montell (Wang et al., 2007 (link)). Generated strains include: Bx-GAL4;Bc¹, Bx-GAL4;tub GAL80^ts, Bx-Gal4;santa-maria¹,Bx-Gal4;;hs-santa-maria, santa-maria¹;UAS-Ras^V12, Bx-Gal4;Df(2L)ED479/CyO,GFP and Bx-Gal4;Df(2L)Exel7031/CyO,GFP. All stocks were maintained on a standard potatomash/molasses medium at 25°C.

Hauling T., Krautz R., Markus R., Volkenhoff A., Kucerova L, & Theopold U. (2014). A Drosophila immune response against Ras-induced overgrowth. Biology Open, 3(4), 250-260.

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Drosophila Genetic Stocks for Functional Studies

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The following stocks were obtained from the Bloomington Stock Center: Oregon R, w¹¹¹⁸, msn-lacZ (msn⁰⁶⁹⁴⁶), puc-lacZ, UAS-Jra, UAS-hep, e16E-GAL4, hs-GAL4, UAS-cdc37-RNAi (JF03184, GD14633, HMS01401), UAS-bsk^DN, UAS-luciferase RNAi, UAS-GFP.nls, UAS-Dcr-2, UAS-cdk2-RNAi (HMS00174; Sopko et al., 2014 ), UAS-cycE-RNAi (HMS00060), and UAS-ckIIα^DN (Lin et al., 2002) . The following stocks were obtained from the Vienna Drosophila RNAi Center in Austria: UAS-cdc37-RNAi (KK102575), UAS-aurora B-RNAi (GD11982, KK112558; Bell et al., 2015 (link)), and UAS-cdc2-RNAi (106130). UAS-cdc37-RNAi (12019R-2) was obtained from the National Institute of Genetics in Japan. D. mojavensis was obtained from The National Drosophila Species Stock Center.

Lee C.W., Kwon Y.C., Lee Y., Park M.Y, & Choe K.M. (2019). cdc37 is essential for JNK pathway activation and wound closure in Drosophila. Molecular Biology of the Cell, 30(21), 2651-2658.

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Drosophila Genetic Mutants and Transgenic Lines

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The following mutant alleles and transgenic lines were used in this study: Girdin² [3 (link)], lgl⁴ [66 (link)], yrt⁷⁵ [27 (link)], UAS-GFP.nls (Bloomington Drosophila Stock Center [BDSC] Stock No. 4776), and UAS-FLAG-Girdin [3 (link)]. Germ line clone females were produced using the FLP-DFS technique [67 (link)], and were used to produce maternal and zygotic (M/Z) mutant embryos. Maternal knockdown of aPKC and lgl was achieved by crossing the driver line matαtub67;15 (obtained from D. St-Johnston, University of Cambridge, Cambridge, UK) to a line containing an inducible shRNA directed against aPKC (BDSC Stock No. 34332) or lgl (BDSC Stock No. 35773) at 25°C.

Biehler C., Wang L.T., Sévigny M., Jetté A., Gamblin C.L., Catterall R., Houssin E., McCaffrey L, & Laprise P. (2020). Girdin is a component of the lateral polarity protein network restricting cell dissemination. PLoS Genetics, 16(3), e1008674.

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Chromocenter Disruption in Drosophila Testes

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All fly stocks were raised on standard Bloomington medium at 25°C. The following fly stocks were used: D1^EY05004(BDSC17340), Df(3R)BSC666 (BDSC26518), UAS-Omi^RNAi (BDSC55165), UAS-GFP-nls (BDSC4776) and UAS-GFP-ER-SR (BDSC59042) were obtained from the Bloomington Drosophila stock center. D1^LL03310 (DGRC140754) and Df31-GFP (DGRC110806) were obtained from the Kyoto stock center. nos-gal4 (Van Doren et al., 1998 (link)), hs-flp;nos-FRT-stop-FRT-gal4,UAS-GFP (Salzmann et al., 2013 (link)), UAS-H2A-YFP (Bellaïche et al., 2001 (link)) and B1 LacO (Vazquez et al., 2002 (link)) have been previously described. A stock containing B chromosomes, mtrm¹²⁶ +B (Bauerly et al., 2014 (link)), was a kind gift from Scott Hawley. Chromocenter disruption was scored in Drosophila testes by assessing {AATAT}_n morphology in GFP +cells that were generated as follows in control (hs-flp; nos-FRT-stop-FRT-gal4,UAS-GFP) and D1 mutant (hs-flp;nos-FRT-stop-FRT-gal4, UAS-GFP;D1^LL03310/Df) flies. Testes were dissected 24 hr following a 20 min heat shock at 37°C. Chromocenters were considered disrupted in Drosophila and mouse when satellite DNA adopted thread-like morphology in interphase nuclei. Micronuclei were scored in 0-3 d testes where early germ cell chromosomes were labeled with H2A-YFP. The genotypes used were, control – nos > H2 A-YFP and D1 mutant – nos > H2 A-YFP; D1LL03310/Df.

Jagannathan M., Cummings R, & Yamashita Y.M. (2018). A conserved function for pericentromeric satellite DNA. eLife, 7, e34122.

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Drosophila Genetic Manipulation Protocol

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The following fly stocks were obtained from the Bloomington Drosophila Stock Center (BDSC): w¹¹¹⁸, UAS-GFP.nls (#4775) and dTet-Gal4 [24 ]. Flies crosses were performed on standard cornmeal-agar medium at 25°C.

Ismail J.N., Mantash S., Hallal M., Jabado N., Khoueiry P, & Shirinian M. (2023). Phenotypic and transcriptomic impact of expressing mammalian TET2 in the Drosophila melanogaster model. Epigenetics, 18(1), 2192375.

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Quantifying GABAergic Neuron Density in Drosophila

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To examine the cell number and organization of GABAergic neurons, a nucleus-targeted UAS-GFP.nls (Bloomington) transgene driven by GAD-Gal4 was introduced into both control and dfmr1 null backgrounds. Brains from 1 day old adult animals were dissected and processed following the immunocytochemical methods described above. Confocal imaging was done of whole brains, with a particular focus on the posterior aspect of the MB calyx region. Maximum intensity projections were generated from Z-stacks to quantify GFP-positive nuclei within the area adjacent to the F-actin delineated MB calyx. From the center of the calyx, a 50μm diameter circle was drawn and all marked nuclear profiles in this brain region counted in control and dfmr1 null animals.

Gatto C.L., Pereira D, & Broadie K. (2014). GABAergic circuit dysfunction in the Drosophila Fragile X syndrome model. Neurobiology of disease, 65, 142-159.

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Fly Circadian Rhythm Manipulation

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Flies were raised on cornmeal-molasses medium in incubators held at 25°C and running on a 12:12 light-dark (LD) schedule. Pdf-GAL4 (FBti0027914), to-GAL4 (FBti0202314), tim-GAL4 (FBti0017922) and cyc⁰¹ were provided by Amita Sehgal. UAS-sgg^Y214F (a hypomorphic sgg allele) (FBti0026626) and UAS-dbt^L (FBti0202311) were provided by Orie Shafer. UAS-GFPnls (FBti0012492), UAS-cyc^DN (FBti0145085), UAS-Cas9.P2 (FBti0166500), tub>GAL80> (FBti0147582), and >stop>GAL80 (FBti0147584) were ordered from the Bloomington Drosophila Stock Center. UAS-sgRNA-per^4x, UAS-sgRNA-tim^3x and UAS-sgRNA-acp^4x were provided by Mimi Shirasu-Hiza. Otd-nls:FLPo (FBtp0093566) was provided by David Anderson. Lsp3.1-GAL4 (FBti0210031) was provided by Brigitte Dauwalder.

Fulgham C.V., Dreyer A.P., Nasseri A., Miller A.N., Love J., Martin M.M., Jabr D.A., Saurabh S, & Cavanaugh D.J. (2021). Central and Peripheral Clock Control of Circadian Feeding Rhythms. Journal of biological rhythms, 36(6), 548-566.

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Drosophila Genetics: Analyzing prod Function

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All fly stocks were raised on standard Bloomington medium at 25°C. The following fly stocks were used: prod^U (BDSC42686), UAS-GFP-nls (BDSC4776), tub-gal4 (BDSC5128), hs-flp (BDSC6), FRT42D, Ubi-nls-GFP (BDSC5626), UAS-dcr-2 (BDSC24650), D1-GFP (BDSC50850), HP1-RFP (BDSC30562) and UAS-DIAP1 (BDSC6657) were obtained from the Bloomington Drosophila stock center. D1^LL03310 (DGRC140754) and FRT42D prod^k08810 (DGRC111248) were obtained from the Kyoto stock center. UAS-prod^RNAi (VDRCv106593) was obtained from the Vienna Drosophila stock center. nos-gal4 and bam-gal4 have been previously described (Chen and McKearin, 2003 (link); Van Doren et al., 1998 (link)). wor-gal4 was a kind gift from Cheng-Yu Lee. Prod-null clones (indicated by loss of GFP signal) were generated as follows – Testes from flies of the genotype hs-flp; FRT42D, Ubi-nls-GFP/FRT42D, prod^k08810 were dissected 48 hr following a 1 hr heat shock at 37°C. For embryo and larval development analysis, flies laid eggs on apple-agar at RT and development was assessed every 24 hr.

Jagannathan M., Cummings R, & Yamashita Y.M. (2019). The modular mechanism of chromocenter formation in Drosophila. eLife, 8, e43938.

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Immunostaining Fly Brain Samples

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Five to seven day old fly brains were dissected in 1% paraformaldehyde in S2 medium, and processed according to a published protocol (Jenett et al., 2012 (link)). Briefly, brains were incubated with the primary antibodies for 3 hours at room temperature and at 4^○C overnight, and with the secondary antibodies for 3 hours at room temperature and 4 days at 4^○C. Incubations were performed in blocking serum (3% normal goat serum). After the final incubation and washes, brains were mounted in vectashield media for imaging. Immunostaining experiments were carried out with a UAS-GFP.nls (Bloomington #4776), which contains a nuclear localization signal but in neurons provides strong labeling of nuclei, somata, and neuronal processes. Antibodies used were rabbit anti- GFP (1:1000, Invitrogen), mouse anti-nc82 (1:50, DSHB), mouse anti-TH (1:200, Chemicon), goat anti-rabbit IgG and goat anti-mouse IgG (1:800, Alexa 488 or Alexa 633 respectively, Invitrogen). Images were obtained using Leica TCS SP8 confocal microscope.

Boto T., Stahl A., Zhang X., Louis T, & Tomchik S.M. (2019). Independent Contributions of Discrete Dopaminergic Circuits to Cellular Plasticity, Memory Strength, and Valence in Drosophila. Cell reports, 27(7), 2014-2021.e2.

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Drosophila Genetic Toolkit Protocols

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We used traffic jam (tj)-Gal4 (Kyoto Stock Center-104055); UAS-GFP^nls (Bloomington Drosophila Stock Center (BDSC-4775); UAS-chinmo^HMS00036-RNA nterference (RNAi) (BDSC-33638); UAS-Dcr-2 (BDSC-24651); mirr-lacZ^cre2 (BDSC-10880); chinmo^ST (Ma et al. 2014 (link)); and the protein trap scrib-GFP (scrib^CA07683) (Buszczak et al. 2007 (link); a gift from Ronald Davis, Scripps Research Florida, FL, USA). We maintained crosses at 25°C in a 12-h light/dark incubator.
Flies were reared on food made with these ingredients: 1800 mL Molasses (LabScientific, Catalog no. FLY-8008-16), 266 g Agar (Mooragar, Catalog no. 41004), 1800 g Cornmeal (LabScientific, Catalog no. FLY-8010-20), 744 g Yeast (LabScientific, Catalog no. FLY-8040-20F), 47 L Water, 56 g Tegosept (Sigma no. H3647-1KG), 560 mL Reagent Alcohol (Fisher no. A962P4), and 190 mL Propionic Acid (Fisher no. A258500).

Grmai L., Harsh S., Lu S., Korman A., Deb I.B, & Bach E.A. (2021). Transcriptomic analysis of feminizing somatic stem cells in the Drosophila testis reveals putative downstream effectors of the transcription factor Chinmo. G3: Genes|Genomes|Genetics, 11(4), jkab067.

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