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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Journal ArticleDOI
GIPC proteins negatively modulate Plexind1 signaling during vascular development.
Jorge Carretero-Ortega,Zinal Chhangawala,Shane Hunt,Carlos Narvaez,Javier Menéndez-González,Tomasz Zygmunt,Xiaochun Li,Jesús Torres-Vázquez +7 more
TL;DR: These findings expand the vascular roles of GIPCs beyond those of the Vascular Endothelial Growth Factor (VEGF)-dependent, proangiogenic GIPC1-Neuropilin 1 complex, recasting GIPPs as negative modulators of antiangiogens PLXND1 signaling and suggest that PLxND1 trafficking shapes vascular development.
Posted ContentDOI
Ancestral circuits for vertebrate colour vision emerge at the first retinal synapse
Takeshi Yoshimatsu,Philipp Bartel,Cornelius Schröder,F. Janiak,François St-Pierre,François St-Pierre,Philipp Berens,Tom Baden,Tom Baden +8 more
TL;DR: In the case of larval zebrafish, this paper showed that the synaptic outputs of red and green cones efficiently rotate the encoding of natural daylight in a principal component analysis (PCA)-like manner to yield primary achromatic and spectrally-opponent axes, respectively.
Journal ArticleDOI
Genetic Deletion of Zebrafish Rab28 Causes Defective Outer Segment Shedding, but Not Retinal Degeneration.
Stephen P Carter,Ailis Moran,David Matallanas,Gavin J. McManus,Oliver E. Blacque,Breandán N. Kennedy +5 more
TL;DR: Cone photoreceptor-specific transgenic reporter lines show Rab28 localizes almost exclusively to outer segments, independently of GTP/GDP nucleotide binding, and co-immunoprecipitation analysis demonstrates tagged Rab28 interacts with components of the phototransduction cascade, including opsins, phosphodiesterase 6C and guanylate cyclase 2D.
Journal ArticleDOI
High Resolution Imaging of DNA Methylation Dynamics using a Zebrafish Reporter.
TL;DR: A transgenic zebrafish reporter of DNA methylation (zebraRDM) via knockin of an mCherry-fused methyl-CpG binding domain (MBD) probe driven by the bactin2 promoter serves as a convenient and powerful tool for high-resolution investigation of methylation dynamics in live animals.
Journal ArticleDOI
Microbial recognition regulates intestinal epithelial growth in homeostasis and disease.
Meghan Ferguson,Edan Foley +1 more
TL;DR: In this paper, the importance of microbial recognition in homeostatic intestinal growth and discuss how immune pathways directly impact stem cell and crypt dynamics, and compare how microbes and subsequent recognition by the intestine influences barrier integrity, intestinal repair and tumorigenesis in Drosophila, zebrafish, mice, and organoids.
References
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Journal ArticleDOI
Stages of embryonic development of the zebrafish.
TL;DR: A series of stages for development of the embryo of the zebrafish, Danio (Brachydanio) rerio is described, providing for flexibility and continued evolution of the staging series as the authors learn more about development in this species.
Journal ArticleDOI
Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein.
Nathan C. Shaner,Robert E. Campbell,Robert E. Campbell,Paul Steinbach,Ben N G Giepmans,Amy E. Palmer,Roger Y. Tsien +6 more
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.
Journal ArticleDOI
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.