Journal ArticleDOI
The Tol2kit: a multisite gateway-based construction kit for Tol2 transposon transgenesis constructs.
Kristen M. Kwan,Esther Fujimoto,Clemens Grabher,Benjamin D. Mangum,Melissa Hardy,Douglas Simon Campbell,John M. Parant,H. Joseph Yost,John P. Kanki,Chi Bin Chien +9 more
TLDR
The Tol2kit greatly facilitates zebrafish transgenesis, simplifies the sharing of clones, and enables large‐scale projects testing the functions of libraries of regulatory or coding sequences.Abstract:
Transgenesis is an important tool for assessing gene function. In zebrafish, transgenesis has suffered from three problems: the labor of building complex expression constructs using conventional subcloning; low transgenesis efficiency, leading to mosaicism in transient transgenics and infrequent germline incorporation; and difficulty in identifying germline integrations unless using a fluorescent marker transgene. The Tol2kit system uses site-specific recombination-based cloning (multisite Gateway technology) to allow quick, modular assembly of [promoter]-[coding sequence]-[3' tag] constructs in a Tol2 transposon backbone. It includes a destination vector with a cmlc2:EGFP (enhanced green fluorescent protein) transgenesis marker and a variety of widely useful entry clones, including hsp70 and beta-actin promoters; cytoplasmic, nuclear, and membrane-localized fluorescent proteins; and internal ribosome entry sequence-driven EGFP cassettes for bicistronic expression. The Tol2kit greatly facilitates zebrafish transgenesis, simplifies the sharing of clones, and enables large-scale projects testing the functions of libraries of regulatory or coding sequences.read more
Citations
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BMP, Wnt and FGF signals are integrated through evolutionarily conserved enhancers to achieve robust expression of Pax3 and Zic genes at the zebrafish neural plate border
TL;DR: This work uses heat-shock overexpression and chemical inhibitors to determine whether, and when, BMP, Wnt and FGF signaling are needed for expression of the NPB specifiers pax3a and zic3 in zebrafish and reveals how BMPs, FGFs and Wnts act cooperatively and redundantly through partially redundant enhancers to achieve robust, specific gene expression in the zebra fish NPB.
Journal ArticleDOI
Bax, Bcl2, and p53 differentially regulate neomycin- and gentamicin-induced hair cell death in the zebrafish lateral line.
TL;DR: A combination of pharmacologic and genetic manipulations are used to assess the contributions of p53, Bax, and Bcl2 in the death of zebrafish lateral line hair cells to suggest a role for mitochondrial p53 activity in promoting hair cell death due to aminoglycosides.
Journal ArticleDOI
Zebrafish as an animal model for biomedical research
TL;DR: Zebrafish have several advantages compared to other vertebrate models used in modeling human diseases, particularly for large-scale genetic mutant and therapeutic compound screenings, and other biomedical research applications.
Journal ArticleDOI
Collective mesendoderm migration relies on an intrinsic directionality signal transmitted through cell contacts.
Julien G. Dumortier,Sandro Martin,Sandro Martin,Dirk Meyer,Frédéric M. Rosa,Frédéric M. Rosa,Nicolas B. David,Nicolas B. David +7 more
TL;DR: An original model of collective migration in which directional information is contained within the moving group rather than provided by extrinsic cues, and constantly maintained in cells by contacts with their neighbors is lead to.
Journal ArticleDOI
Genetic inducible fate mapping in larval zebrafish reveals origins of adult insulin-producing β-cells
TL;DR: It is shown that the PNCs represent a population of progenitors that can differentiate to multiple lineages, including adult ductal cells, centroacinar cells (CACs) and endocrine cells, and CACs are more closely related toendocrine cells by lineage as they share a common progenitor.
References
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Journal ArticleDOI
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TL;DR: The latest red version matures more completely, is more tolerant of N-terminal fusions and is over tenfold more photostable than mRFP1, and three monomers with distinguishable hues from yellow-orange to red-orange have higher quantum efficiencies.
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An Hsp70-like protein in the ER: identity with the 78 kd glucose-regulated protein and immunoglobulin heavy chain binding protein.
Sean Munro,Hugh R.B. Pelham +1 more
TL;DR: A cDNA clone is characterized that encodes a protein related to the 70 kd heat shock protein, but is expressed in normal rat liver, and it is identical with two previously described proteins: GRP78, whose synthesis is induced by glucose starvation, and BiP, which is found bound to immunoglobulin heavy chains in pre-B cells.
Journal ArticleDOI
GAL4-VP16 is an unusually potent transcriptional activator
TL;DR: It is shown that the hybrid protein (GAL4-VP16) activates transcription unusually efficiently in mammalian cells when bound close to, or at large distances from the gene, and suggested that the activating region of VP16 may be near-maximally potent.
Journal ArticleDOI
DNA Cloning Using In Vitro Site-Specific Recombination
TL;DR: A method called recombinational cloning is described that uses in vitro site-specific recombination to accomplish the directional cloning of PCR products and the subsequent automatic subcloning of the DNA segment into new vector backbones at high efficiency.