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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Functional Investigation of a Non-coding Variant Associated with Adolescent Idiopathic Scoliosis in Zebrafish: Elevated Expression of the Ladybird Homeobox Gene Causes Body Axis Deformation.
Long Guo,Hiroshi Yamashita,Ikuyo Kou,Aki Takimoto,Makiko Meguro-Horike,Shin-ichi Horike,Tetsushi Sakuma,Shigenori Miura,Taiji Adachi,Takashi Yamamoto,Shiro Ikegawa,Yuji Hiraki,Chisa Shukunami,Chisa Shukunami +13 more
TL;DR: This study presents a novel pathological feature of LBX1 and its zebrafish homologs in body axis deformation at various stages of embryonic and subsequent growth in zebra fish.
Journal ArticleDOI
The feelgood mutation in zebrafish dysregulates COPII-dependent secretion of select extracellular matrix proteins in skeletal morphogenesis.
David B. Melville,Mercedes Montero-Balaguer,Daniel S. Levic,Kevin M. Bradley,Jeffrey R. Smith,Antonis K. Hatzopoulos,Ela W. Knapik +6 more
TL;DR: Results indicate that Creb3l2 modulates the availability of COPII machinery in a tissue- and cargo-specific manner, which could lead to a better understanding of the etiology of human craniofacial and skeletal birth defects as well as adult-onset diseases that are linked to dysregulated ECM deposition, such as arthritis, fibrosis or osteoporosis.
Journal ArticleDOI
Vertebrate epidermal cells are broad-specificity phagocytes that clear sensory axon debris.
TL;DR: The authors found that epidermal cells rapidly internalize axon debris into dynamic phosphatidylinositol 3-monophosphate-positive phagosomes that mature into phagolysosomes using a pathway similar to professional phagocytes.
Journal ArticleDOI
In vivo labeling of zebrafish motor neurons using an mnx1 enhancer and Gal4/UAS
TL;DR: The use of the 125‐bp mnx1 promoter for transient transgenic expression or for the generation of stable transgenic fish lines will facilitate the study of motor neuron development and neurodegenerative processes.
Journal ArticleDOI
EphA4 and EfnB2a Maintain Rhombomere Coherence by Independently Regulating Intercalation of Progenitor Cells in the Zebrafish Neural Keel
TL;DR: A model in which Eph and Efn-dependent cell affinity within rhombomeres serve to maintainrhombomere organization during the potentially disruptive process of teleost neurulation is proposed.
References
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Journal ArticleDOI
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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.