Section (A–A′″) and section (B–B′″) are the same section, focusing on different cell groups when imaging. 1C, arrow). Signals were visualized on a Nikon Eclipse E3600 stereomicroscope with filters for both fluorescent stains. Isolation of a Zebrafish Rod Opsin Promoter to Generate a Transgenic Zebrafish Line Expressing Enhanced Green Fluorescent Protein in Rod Photoreceptors* Received for publication, November 20, 2000, and in revised form, January 12, 2001 Published, JBC Papers in Press, January 18, 2001, DOI 10.1074/jbc.M010490200 MPTP (1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine) treatments induced a modest but significant loss of DA neurons in groups 2–6 of the vDC. Green fluorescent protein (GFP) as a marker of aryl hydrocarbon receptor (AhR) function in developing Zebrafish (Danio rerio). Delayed effects of methylmercury on the mitochondria of dopaminergic neurons and developmental toxicity in zebrafish larvae (Danio rerio). Zebrafish as a model system for mitochondrial biology and diseases. A 27‐kb fragment (including approximately 13 kb of dat 5′‐flanking region) was then cloned into a pGEM‐Tol2 vector in another round of homologous recombination. In our recent screening of the distribution patterns of fluorescent compounds in live zebrafish larvae, fifteen compounds with tissue-specific distributions were identified. 10.1002/(SICI)1520-6408(1999)25:2<158::AID-DVG10>3.0.CO;2-6. Kisspeptin-1 regulates forebrain dopaminergic neurons in the zebrafish. Figure 1: (A) Zebrafish embryo under a bright field microscope5. This autofluorescence may obscure all but exceptionally bright specific labels when viewed by widefield epifluorescence. More than 90% of embryos injected with this construct in the presence of tol2 transposase mRNA showed GFP expression in areas located roughly between the eyes at 2 days post‐fertilization (dpf). Transgenic zebrafish embryos expressing green fluorescent protein in the developing notocord and ventral neural tube. doi: 10.1002/(SICI)1520-6408(1999)25:2<158::AID-DVG10>3.0.CO;2-6. Embryos for whole mount in situ hybridization and immunostaining were fixed in 4% paraformaldehyde (PFA) in phosphate‐buffered saline (PBS), then dehydrated in 100% methanol, and stored at −20°C in 100% methanol until use. 2011;6(5):e20654. Epub 2013 Mar 11. Tg(dat:EGFP) embryos were untreated (Ctrl) or treated with MPTP at 1 mM from 24 hours post‐fertilization (hpf), and examined under fluorescence microscope at 3 days post‐fertilization (dpf) (A,B) and 5 dpf (C,D). The total size of the inserted DNA is 27 kb, containing the whole dat genomic sequence except for the last coding exon and 3′‐flanking region. Zebrafish transgenics have also been widely used to study gene regulation, an example of which is the analysis of the multiple enhancers of the DFN3 ortholog pou3f4 (Robert-Moreno et al., 2010). Dev. Arrows show GFP/TH‐positive cells in the retina. USA.gov. A zebrafish cDNA encoding a novel keratin protein was characterized and named keratin8, or krt8. GloFish® fluorescent fish come in a variety of species and colors of tropical fish. As GFP starts being expressed at around 20 hpf in Tg(dat:EGFP) embryos, we exposed embryos, starting at 24 hpf, to different concentrations of MPTP (100 μM, 500 μM and 1 mM). As a teleost class of fish species, zebrafish … 4B–D′″). National Center for Biotechnology Information, Unable to load your collection due to an error, Unable to load your delegates due to an error. Bringing Color to Life! 3 A … Zebrafish were the first GloFish available in pet stores, and are now sold in bright red, green, orange-yellow, blue, pink, and purple fluorescent colors. Next, we used ZF-Mapper to analyze the fluorescent images of cancer xenograft zebrafish. Research output: … Washington University in St. Louis is now home to one of the largest zebrafish facilities in the world. In this article, we’ll explore fluorescent transgenic zebrafish lines in particular, and how they can be used in Drug Discovery and development. Epub 2012 Nov 28. This included the DA neurons in the vDC (groups 2–6). Arrows in B indicate the boundaries of scales dividing the krt8‐expressing and nonexpressing portions. Additional sites of ectopic GFP expression included the jaw and groups of cells at or near the midbrain–hindbrain boundary. 2013 May;55(4):422-33. doi: 10.1111/dgd.12042. Print 2017 Jun 30. A 1905 base pair region of the human CYP1A1 promoter/enhancer region was regulated by AhR in zebrafish liver cells after exposure to TCDD (10 nM) in a transient transfection assay. An adult zebrafish brain showing fluorescent granular perithelial cells (green) atop blood vessels (purple). (2010). The 2.2-kb ck promoter Scale bars = 100 μm in A–A″,B–B″,C–C″,D–D″,E–E″; 25 μm in A′″,B′″,C′″,D′″,E′″.  |  EGFP expression was detected in the ventral-nasal eye at 3 days postfertilization and spread throughout the eye. Double fluorescent in situ hybridization with GFP and dat cRNA probes indicates that the transgene is expressed as predicted in dat‐expressing cells, notably in the ventral diencephalon (Fig. GFP‐positive individuals were raised to sexual maturity and outcrossed with wild type adult fish to identify transgenic carriers. … Data were expressed as mean ± SD. The green fluorescence comes from a sonic hedgehog-GFP reporter transgene acting in these living embryos. The 2.2-kb ck promoter was sufficient to direct GFP expression in skin epithelia, although a weak expression in muscle was also observed in a few embryos. Guoying Wang. Scale bar = 100 μm. The arrow indicates a group of cells in the hindbrain that express GFP and may correspond to some of the dat‐expressing cells reported by Holzschuh et al. Characterization of transgenic zebrafish lines that express GFP in the retina, pineal gland, olfactory bulb, hatching gland, and optic tectum. Arrowheads show GFP/TH‐positive cells in the Ob, Pr and Hc. Faithful expression of living color reporter genes in transgenic medaka under two tissue-specific zebrafish promoters. (1995). The antimicrobial and antioxidant properties of garagurt: traditional Cornelian cherry (Cornus mas) marmalade. 109, No. Whole‐mount immunostaining for green fluorescent protein (GFP) and tyrosine hydroxylase (TH) in 3 days postfertilization (dpf) Tg(dat:EGFP) larvae. Organization and projection pattern of dopaminergic neurons in the diencephalon, The relationship between dlx and gad1 expression indicates highly conserved genetic pathways in the zebrafish forebrain, Neuroprotection of MPTP‐induced toxicity in zebrafish dopaminergic neurons, Targeting retinal dopaminergic neurons in tyrosine hydroxylase‐driven green fluorescent protein transgenic zebrafish, The parkinsonian toxin 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP): a technical review of its utility and safety, The teleostean (zebrafish) dopaminergic system ascending to the subpallium (striatum) is located in the basal diencephalon (posterior tuberculum), Development of the catecholaminergic system in the early zebrafish brain: an immunohistochemical study, Orthopedia homeodomain protein is essential for diencephalic dopaminergic neuron development, MPTP and MPP+ target specific aminergic cell populations in larval zebrafish, Comprehensive catecholaminergic projectome analysis reveal single‐neuron integration of zebrafish ascending and descending dopaminergic systems, Visualization of monoaminergic neurons and neurotoxicity of MPTP in live transgenic zebrafish, Impaired dopaminergic neuron development and locomotor function in zebrafish with loss of pink1 function, Two tyrosine hydroxylase genes in vertebrates New dopaminergic territories revealed in the zebrafish brain, Differential expression of dopaminergic cell markers in the adult zebrafish forebrain. 1B and data not shown). The third clone, expressed ubiquitously in all tissues, is derived from an acidic ribosomal phosphoprotein P0 (arp) gene. 6). Our experiments, therefore, further demonstrated that zebrafish embryos can faithfully express exogenously introduced genes under the control of zebrafish promoters. The following abbreviations are used: olfactory bulb (Ob), pretectum (Pr), ventral diencephalon (vDC), amacrine cells (Ac) and caudal hypothalamus (Hc). In the present study, three different zebrafish cDNA clones were isolated and sequenced completely, and their expression patterns were characterized by whole-mount in situ hybridization as well as by Northern blot hybridization. Here, we test whether LNPs can be used to deliver a reporter green fluorescent protein (gfp) mRNA to different tissues in zebrafish embryos. Here, we describe the development of a novel zebrafish line which uses the putative promoter of Myelin Protein Zero (mpz), a major structural protein in myelin, to drive expression of Enhanced Green Fluorescent Protein (mEGFP) specifically in the processes and nascent internodes of myelinating glia. The zebrafish has proven to be an excellent vertebrate model for developmental biology and the study of human diseases (Driever et al., 1994, 1996; Zon, 1999). Recently "Electric Green", "Sunburst Orange", "Moonrise Pink", "Starfire Red", "Cosmic Blue", and "Galactic Purple" colored tetra (Gymnocorymbus ternetzi), an "Electric Green" tiger barb (Punt… Embryos were staged as hours (hpf) or days (dpf) post‐fertilization according to specific morphological features outlined by Kimmel et al. In Tg(dat:EGFP) fish, all major clusters of DA neurons are correctly labeled with GFP during early embryogenesis, including those in the vDC. A 1.2-kbp promoter fragment was cloned upstream of the enhanced green fluorescent protein (EGFP) cDNA and microinjected into 1- to 2-cell stage zebrafish embryos. The Aequorea victoria green fluorescent protein can be used as a reporter in live zebrafish embryos. Experimental Models to Study Autism Spectrum Disorders: hiPSCs, Rodents and Zebrafish. 2005). Although the zebrafish has become a popular model organism for vertebrate developmental and genetic analyses, its use in transgenic studies still suffers from the scarcity of homologous gene promoters. A–D: Sagittal sections of an adult zebrafish hybridized with the krt8 antisense riboprobe. 1A, Liu et al., 2003). Differential gene expression following TLR stimulation in rag1-/- mutant zebrafish tissues and morphological descriptions of lymphocyte-like cell populations. This allows us to view the migration of the axons in … Green Fluorescent Protein (GFP) is a common protein used to localise proteins, observe protein interactions and quantify gene expression. Although several genes have been found to be associated with familial inheritable PD, the detailed etiology of PD is still not well understood (Cookson, 2005). In this video the motor neurons in a transgenic zebrafish embryo have been labelled with Green Fluorescent protein (GFP). Selective toxicity of L-DOPA to dopamine transporter-expressing neurons and locomotor behavior in zebrafish larvae. Overall, the Tg(dat:EGFP) transgenic fish may be used as a live animal model for to better understand the mechanisms of DA neuron development and as a model to study pathological mechanisms associated with Parkinson's disease. Embryos were obtained from an outcross between the dat:EGFP transgenic fish and wild‐type individuals. Biosci Rep. 2017 Jun 8;37(3):BSR20170199. All zebrafish husbandry were performed in accordance with institutional, national ethical, and animal welfare guidelines. Zebrafish Neuromast Labeling Protocols 12/2/05 Working with fixed zebrafish. This regulatory region was fused to the cDNA sequence encoding green fluorescent protein (GFP) of jellyfish (Aequorea victoria). All experiments were independently repeated at least three times. doi: 10.1371/journal.pone.0184077. The GFP probe was labeled with DNP and revealed with tyr‐Cy3. (2003). In this study, we took advantage of the fact that DAT expression distinguishes DA neurons from other catecholaminergic (CA) neurons in the developing embryo (Holzschuh et al., 2001) and produced a line of transgenic fish in which the green fluorescent protein (GFP) is expressed under the control of regulatory elements of the zebrafish dat gene. Faithful expression of green fluorescent protein (GFP) in transgenic zebrafish embryos under control of zebrafish gene promoters. This process is experimental and the keywords may be updated as the learning algorithm improves. Embryos and larvae for double immunohistochemistry were fixed in 4% PFA/1× PBS overnight at 4°C, rinsed in 1× PBS and placed in 30% sucrose/1× PBS to equilibrate. We show that LNP-packaged gfp mRNA can be delivered, through injection, and taken up by cells in multiple tissues in zebrafish embryos without any apparent detrimental effects on embryonic health or survival. All sections are shown as anterior to the left. 4). Double immunostaining for green fluorescent protein (GFP) and tyrosine hydroxylase (TH) on transverse cryosections of Tg(dat:EGFP) larvae at 3 days postfertilization (dpf). Advantages of the Zebrafish Platform in Drug Screening. The tyrosine hydroxylase (th) and dopamine transporter (dat) genes are commonly used as markers for mature DA neurons. many times, GloFish® fluorescent zebra fish can stay in a quite huge temperature selection, everywhere from sixty 4-86°F (18–30°C), yet want temperatures of seventy two-80°F (22-27°C). The 5 dpf Tg(dat:EGFP) embryos were horizontally cryosectioned and stained with anti‐GFP and anti‐TH antibodies. The 3 dpf Tg(dat:EGFP) larvae were stained with anti‐GFP and anti‐TH antibodies, followed by confocal microscopy. Here, we re‐examined this issue using the Tg(dat:EGFP) fish. The 1.5-kb mck promoter/gfp was expressed exclusively in skeletal muscles and not elsewhere. Zebrafish is also amenable to genetics due to its relatively short generation time (2–3 months). The embryonic zebrafish is a nearly ideal model system in which to use time-lapse imaging to study the development of the vertebrate nervous system in vivo. Color:Green Fluorescent GloFish aquarium kits, lighting and décor create an underwater fluorescent wonderland that appeals to all ages and levels of expertise. Working off-campus? These keywords were added by machine and not by the authors. In order to test the fidelity of zebrafish embryos in transgenic expression, the promoters of the three genes were isolated using a rapid linker-mediated PCR approach and subsequently ligated to a modified green fluorescent protein (gfp) reporter gene. EGFP (in green) was inserted in frame at the beginning of exon 1. (2010). Research output: Contribution to journal › Article › peer-review 8A–D). 2003 May;227(1):14-26. doi: 10.1002/dvdy.10273. Injected embryos were screened for GFP expression in the desired area between 48 and 72 hpf. Dopaminergic (DA) neurons synthesize dopamine from tyrosine through the action of tyrosine hydroxylase (TH), the first and rate‐limiting enzyme, and DOPA decarboxylase. 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Was accomplished by fusing GFP sequences to Islet-1 promoter/enhancer sequences that were sufficient for neural-specific expression embryos at 3.... Melanoma cells labeled green fluorescent zebrafish the krt8 antisense riboprobe and treatments for dopaminergic.. Confocal microscopy observation were performed in accordance with institutional, national ethical, and Optic tectum other including. Gfp‐Positive DA neurons in green fluorescent zebrafish ventral diencephalon mylz2 promoter M, Searson PC, Lee.! Outcrossed with wild type adult fish to identify transgenic carriers, focusing different!

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