Methods for targeted mutagenesis in zebrafish using TALENs

Hwang, Woong Y.; Peterson, Randall T.; Yeh, Jing-Ruey Joanna

doi:10.1016/j.ymeth.2014.04.009

Cited by 30 publications

(19 citation statements)

References 56 publications

(102 reference statements)

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“…The specificity of ZFN targeting results from zinc finger alpha helices that recognize specific three base pair DNA sequences; when several of these helices are tethered together and coupled to the FokI nuclease, DNA double-stranded breaks can be created in a specific sequence of 18–21 bp (Urnov, Rebar, Holmes, Zhang, & Gregory, 2010). The TALEN-targeted mutagenesis technology (transcription activator-like effector nucleases) uses a similar concept to ZFNs with sequence specificity provided by modular protein domains that recognize specific base pair sequences (Huang et al, 2012; reviewed in Hwang, Peterson, & Yeh, 2014). These targeted mutagenesis technologies are also similar in that the double-stranded break which is created is repaired through cellular nonhomologous end joining (NHEJ) mechanisms, resulting in variably sized insertions and deletions (indels), disrupting the genomic sequence of your target GOI.…”

Section: Studying Molecular Requirements Through Loss-of-function mentioning

confidence: 99%

Methods to study maternal regulation of germ cell specification in zebrafish

Kaufman

Marlow

2016

Methods in Cell Biology

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The process by which the germ line is specified in the zebrafish embryo is under the control of maternal gene products that were produced during oogenesis. Zebrafish are highly amenable to microscopic observation of the processes governing maternal germ cell specification because early embryos are transparent, and the germ line is specified rapidly (within 4–5 h post fertilization). Advantages of zebrafish over other models used to study vertebrate germ cell formation include their genetic tractability, the large numbers of progeny, and the easily manipulable genome, all of which make zebrafish an ideal system for studying the genetic regulators and cellular basis of germ cell formation and maintenance. Classical molecular biology techniques, including expression analysis through in situ hybridization and forward genetic screens, have laid the foundation for our understanding of germ cell development in zebrafish. In this chapter, we discuss some of these classic techniques, as well as recent cutting-edge methodologies that have improved our ability to visualize the process of germ cell specification and differentiation, and the tracking of specific molecules involved in these processes. Additionally, we discuss traditional and novel technologies for manipulating the zebrafish genome to identify new components through loss-of-function studies of putative germ cell regulators. Together with the numerous aforementioned advantages of zebrafish as a genetic model for studying development, we believe these new techniques will continue to advance zebrafish to the forefront for investigation of the molecular regulators of germ cell specification and germ line biology.

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Section: Studying Molecular Requirements Through Loss-of-function mentioning

confidence: 99%

Methods to study maternal regulation of germ cell specification in zebrafish

Kaufman

Marlow

2016

Methods in Cell Biology

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“…13 The targeted introduction of mutations using sequencespecific TALENs or the CRISPR system have recently been successfully applied to generate loss-of-function alleles by specifically targeting open reading frames or deletion or inversion of whole-chromosomal regions in vivo with efficiencies in zebrafish similar to those obtained using zinc-finger nucleases and transcriptionlike nucleases. 14 TALENs comprise a non-specific DNA-cleaving nuclease fused to an engineered DNA-binding domain, which contains a series of tandem repeats. Binding of two TALENs to their respective target sites, reconstitutes the active nuclease domain resulting in cleavage of the targeted genomic locus by inducing a double-strand break.…”

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confidence: 99%

The zebrafish eye—a paradigm for investigating human ocular genetics

Richardson¹,

Tracey‐White²,

Webster

et al. 2016

Eye

134

140

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Although human epidemiological and genetic studies are essential to elucidate the aetiology of normal and aberrant ocular development, animal models have provided us with an understanding of the pathogenesis of multiple developmental ocular malformations. Zebrafish eye development displays in depth molecular complexity and stringent spatiotemporal regulation that incorporates developmental contributions of the surface ectoderm, neuroectoderm and head mesenchyme, similar to that seen in humans. For this reason, and due to its genetic tractability, external fertilisation, and early optical clarity, the zebrafish has become an invaluable vertebrate system to investigate human ocular development and disease. Recently, zebrafish have been at the leading edge of preclinical therapy development, with their amenability to genetic manipulation facilitating the generation of robust ocular disease models required for large-scale genetic and drug screening programmes. This review presents an overview of human and zebrafish ocular development, genetic methodologies employed for zebrafish mutagenesis, relevant models of ocular disease, and finally therapeutic approaches, which may have translational leads in the future.

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“…As such, due to the absence of genetic evidence, some of the zebrafish studies highlighted in this review may need to be supplemented by more accurate genetargeting approaches. To this end, the use of sequence-specific DNA-binding endonucleases, such as Zinc finger nucleases (ZFNs) (Foley et al, 2009), transcription activator-like effector nucleases (TALENs) (Hwang et al, 2014) or the CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 system (Varshney et al, 2015) to introduce double-strand breaks at specific loci in the genome can enable generation of mutant embryos.…”

Section: Conclusion/discussionmentioning

confidence: 99%

Etiology of intracerebral hemorrhage (ICH): novel insights from Zebrafish embryos

Eisa-Beygi

Rezaei²

2016

Int. J. Dev. Biol.

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Intracerebral hemorrhage (ICH) is the most severe subtype of stroke. Treatment options are scarce and given the high morbidity and mortality, relatively ineffective. Since patients with ICH may have an unknown heritable component, the need to identify potential risk factors necessitates the use of animal models to elucidate the genetic underpinnings of neurovascular development and, thereby, identify candidate regulatory pathways that are likely to be disrupted in patients with ICH. Zebrafish (Danio rerio) exhibits the anatomical and physiological complexity of a closed circulatory system observed in all vertebrates (with arteries, veins and capillaries). Moreover, studies over the last decade, aided by the application of chemical mutagenesis screens, morpholino mediated knockdown approaches and tissue-specific transgenic markers, have paved the way for the identification of several genes and signaling pathways that regulate developmental neurovascular stabilization. We hypothesize that mutations in these genes or pharmacological perturbations of these gene-products may account, at least in part, for the etiology of some forms of spontaneous ICH in humans.

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Methods for targeted mutagenesis in zebrafish using TALENs

Cited by 30 publications

References 56 publications

Methods to study maternal regulation of germ cell specification in zebrafish

Methods to study maternal regulation of germ cell specification in zebrafish

The zebrafish eye—a paradigm for investigating human ocular genetics

Etiology of intracerebral hemorrhage (ICH): novel insights from Zebrafish embryos

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