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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Differentiated melanocyte cell division occurs in vivo and is promoted by mutations in Mitf
Kerrie L. Taylor,James A. Lister,Zhiqiang Zeng,Hironori Ishizaki,Caroline Anderson,Robert N. Kelsh,Ian J. Jackson,E. Elizabeth Patton +7 more
TL;DR: Results reveal a novel pathway of differentiated melanocyte division in vivo, and suggest Mitf activity is essential for maintaining cell cycle arrest in differentiated melanocytes, and suggests differentiated cell division is relevant to melanoma progression.
Journal ArticleDOI
Motoneurons are essential for vascular pathfinding
TL;DR: Zebrafish Netrin 1a directs Dcc-mediated axon guidance of motoneurons and that this neural guidance function is essential for lymphangiogenesis, providing a definitive example of the requirement of axons in endothelial guidance leading to the parallel patterning of nerves and vessels in vivo.
Journal ArticleDOI
Iroquois Proteins Promote Skeletal Joint Formation by Maintaining Chondrocytes in an Immature State
Amjad Askary,Lindsey Mork,Sandeep Paul,Xinjun He,Audrey K. Izuhara,Suhasni Gopalakrishnan,Justin K. Ichida,Andrew P. McMahon,Sonja Dabizljevic,Rodney M. Dale,Francesca V. Mariani,J. Gage Crump +11 more
TL;DR: It is shown that two members of the Iroquois transcriptional repressor family, Irx7 and Irx5a, function to block chondrocyte maturation at the developing hyoid joint of zebrafish.
Journal ArticleDOI
Genetic and neuronal regulation of sleep by neuropeptide VF
Daniel A. Lee,Andrey Andreev,Thai V. Truong,Audrey Chen,Andrew J. Hill,Grigorios Oikonomou,Uyen Pham,Young Hong,Steven Tran,Laura Glass,Viveca Sapin,Jae Engle,Scott E. Fraser,David A. Prober +13 more
TL;DR: It is reported that the vertebrate hypothalamic RFamide neuropeptide VF (NPVF) regulates sleep in the zebrafish, a diurnal vertebrate and suggested that RFamide Neuropeptides participate in an ancient and central aspect of sleep control.
Journal ArticleDOI
CFTR Protects against Mycobacterium abscessus Infection by Fine-Tuning Host Oxidative Defenses
Audrey Bernut,Audrey Bernut,Christian Dupont,Nikolay V. Ogryzko,Aymeric Neyret,Jean-Louis Herrmann,R. Andres Floto,Stephen A. Renshaw,Laurent Kremer,Laurent Kremer +9 more
TL;DR: A CFTR-depleted zebrafish model is exploited, recapitulating CF immuno-pathogenesis, to study the contribution of CFTR in innate immunity against M. abscessus by mounting effective oxidative responses.
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.