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Open AccessJournal ArticleDOI

Out with the old, in with the new: reassessing morpholino knockdowns in light of genome editing technology.

Stefan Schulte-Merker, +1 more
- 15 Aug 2014 - 
- Vol. 141, Iss: 16, pp 3103-3104
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TLDR
Different approaches to describe how the first description of a loss-of-function phenotype in zebrafish should be accomplished are discussed, with a specific focus on how to describe the effects of morpholino side effects.
Abstract
Morpholino oligomers have been used widely and for many years in the zebrafish community to transiently knock down the function of target genes. It has often been difficult, however, to reliably discriminate between specific and non-specific effects, and thus generally accepted guidelines to control for morpholino side effects do not exist. In light of recent methodologies to generate mutant lines in virtually any zebrafish gene, we discuss these different approaches with a specific focus on how the first description of a loss-of-function phenotype in zebrafish should be accomplished.

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Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)

Daniel J. Klionsky, +2983 more
- 08 Feb 2021 - 
TL;DR: In this article, the authors present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes.
Journal ArticleDOI

Consensus guidelines for the use and interpretation of angiogenesis assays

Patrycja Nowak-Sliwinska, +90 more
- 01 Aug 2018 - 
TL;DR: In vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis are described and critical aspects that are relevant for their execution and proper interpretation are highlighted.
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Rapid reverse genetic screening using CRISPR in zebrafish

TL;DR: It is concluded that CRISPR can be used as a powerful reverse genetic screening strategy in vivo in a vertebrate system and identified two new genes involved in electrical-synapse formation.
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Maximizing mutagenesis with solubilized CRISPR-Cas9 ribonucleoprotein complexes

TL;DR: The results establish that optimally solubilized, in vitro assembled fluorescent Cas9-sgRNA RNPs provide a reproducible reagent for direct and scalable loss-of-function studies and applications beyond zebrafish experiments that require maximal DNA cutting efficiency in vivo.
References
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Journal ArticleDOI

Efficient genome editing in zebrafish using a CRISPR-Cas system

TL;DR: It is shown that the CRISPR-Cas system functions in vivo to induce targeted genetic modifications in zebrafish embryos with efficiencies similar to those obtained using zinc finger nucleases and transcription activator-like effector nucleases.
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Effective targeted gene ‘knockdown’ in zebrafish

TL;DR: It is shown here that antisense, morpholino-modified oligonucleotides (morpholinos) are effective and specific translational inhibitors in zebrafish, and conserved vertebrate processes and diseases are now amenable to a systematic, in vivo, reverse-genetic paradigm using zebra fish embryos.
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Improving CRISPR-Cas nuclease specificity using truncated guide RNAs

TL;DR: It is reported that truncated gRNAs, with shorter regions of target complementarity <20 nucleotides in length, can decrease undesired mutagenesis at some off-target sites by 5,000-fold or more without sacrificing on-target genome editing efficiencies.
Journal ArticleDOI

The identification of genes with unique and essential functions in the development of the zebrafish, Danio rerio

TL;DR: It is estimated that the 372 genes defined by the mutants probably represent more than half of all genes that could have been discovered using the criteria of the screen, and the limits and the potentials of a genetic saturation screen in the zebrafish.
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