applications of zebrafish embryos

Finally, like many other animals, sensory processes guide mating behavior in fish. This observation unlocked novel research into genetic compensation, a previously unknown mechanism of gene regulation (El-Brolosy et al., 2019). To highlight some of the features that make zebrafish useful, I’ll outline two recent lineage tracing approaches that led to important insights in developmental biology. PubMed PMID: 29853675. Please check your Internet connection and reload this page. Then, leave the fish to acclimate to each other overnight. These embryos have the great advantage that they develop as \"see through\" embryos, that is, all internal development can be clearly observed from the outside in the living embryo. Live embryos expressing fluorescent proteins in specific cell types can be mounted and visualized with confocal microscopy, allowing researchers to examine cell movements in developing tissues over time. This study also showed that in the absence of the signaling pathway regulated by one-eyed pinhead, incorrect proportions of otherwise normal cell types are produced, rather than novel, abnormal cell types. The popular UAS/Gal4 and Cre/Lox gene regulatory systems have also been successfully implemented in zebrafish. This post was contributed by Katherine Rogers, a postdoctoral researcher at the Friedrich Miescher Lab of the Max Planck Society. Compared to the opaque mouse embryo, where similar processes occur over days inside the mother, the zebrafish is an ideal system to observe embryogenesis live. In summary, the ZEG device enables rapid genotyping of live zebrafish embryos and larvae, and animals are available for downstream applications, testing, or raising. A common approach to unravel the gene networks that drive biology is to examine the consequences of manipulating genes. For example, specific tissues such as blood cells or neurons can be fluorescently labeled, allowing researchers to observe changes to these tissues under different experimental conditions. The rapid advances being made in gene editing utilizing the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 (CRISPR- associated protein 9) system have led to increasing interest in the application of this technology in zebrafish embryos for gene knockout. Any time you need to move embryos from one dish to another, use a transfer pipet. The exposure of zebrafish embryos was performed according to the exposure scenario described for the fish embryo acute toxicity test with the exception of 10 instead of 5 embryos per glass vial. Zebrafish embryos, used as the species under study in this work, were raised within the acoustic field by the authors, allowing the biological effects on such early zebrafish embryos, at each developmental stage and within the ASWF creating the acoustic levitation (AL) technology used, to be studied. El-Brolosy, Mohamed A., et al. In addition, the Zebrafish Model Organism Database ZFIN actively curates information on mutants and other resources. PubMed PMID: 27443928. Some protocols require a few additional steps for embryo maintenance. Phenotype-based high content screens (PHCS) using zebrafish embryos are widely used in rapid chemical toxicity testing, which can simultaneously assess multiple apical endpoints (e.g., mortality and malformation effects). You’ve just watched JoVE’s video on zebrafish breeding and embryo handling. Citation: Lambert CJ, Freshner BC, Chung A, Stevenson TJ, Bowles DM, Samuel R, et al. The external development of zebrafish embryos allows for genetic manipulation by microinjection of DNA, RNA, or gene knockdown reagents. Zebrafish one to eight-cell embryos were microinjected with 6.5 ng VaDNA together with the firefly and Renilla reporter vectors as indicated in Legend to Figure 2 alone or in combination with 4 ng of the indicated morpholinos. Finally, the transparency of the zebrafish embryos makes them well suited for microscopy. Keller, Philipp J., et al. doi: 10.1371/journal.pone.0043850. Zebrafish are also amenable to manipulation of brain activity: optogenetic approaches can be used to determine the consequences of activating or silencing specific neurons in living larvae. Due to their small size and the relatively simple nature of … ", Felker, A., and C. Mosimann. ", Nüsslein-Volhard, Christiane. Several zebrafish-specific gene expression databases that provide information about when and where genes are active in developing embryos are available. If you have any questions, please do not hesitate to reach out to our customer success team. Circulation begins in the zebrafish embryo at approximately 24 hours post fertilization (hpf). Please check your Internet connection and reload this page. Evaluation of 14 organic solvents and carriers for screening applications in zebrafish embryos and larvae. Epub 2012 Oct … The accessibility of its externally fertilized embryos allows numerous manipulations, and the transparency of the zebrafish embryo permits direct serial visualization of biological processes in vivo (Beis and Stainier, 2006). Academic Press, 2016. Development 139.22 (2012): 4099-4103. The same technique can also be used to make transgenic fish in which specific structures are marked by the expression of fluorescent proteins. To see through this problem, scientists supplement the embryo water with PTU, which inhibits production of the pigment melanin. Next, pour the water and the embryos through a mesh tea strainer for collection. "Pan-neuronal calcium imaging with cellular resolution in freely swimming zebrafish." Let’s go over some pointers. They are important for your experiments! Despite these caveats, several features of zebrafish—including the available genetic resources and transparency of the embryo—have proven useful in a variety of research applications. "Reconstruction of zebrafish early embryonic development by scanned light sheet microscopy." Several different methods have been developed, relying on established reagents such as CRISPR/Cas9, Tol2, and the high-quality sequenced zebrafish genome. These mutants are used extensively to study development and disease. A high-quality sequence assembly of the zebrafish genome was published in 2013, revealing that about 70% of human genes have a counterpart in zebrafish. “Brainbow” fish lines have been developed to enable high-resolution mapping of the brain by uniquely labeling neurons. Periodic mating keeps eggs healthy, but making all those eggs takes a lot of work, so individual females should only be mated once per week. Zebrafish eggs are fertilized and develop outside of the mother's body making visualization with high resolution imaging easier for researchers. Also, their optical transparency facilitates high-resolution imaging of blood vessels in the developing embryo. Zebrafish undergo sexual reproduction, so breeding requires both male and female fish. Posted by Katherine W. Rogers is a postdoctoral researcher in Patrick Müller’s group at the FriedrichMiescher Laboratory of the Max Planck Society in Tübingen, Germany. If you want more info regarding data storage, please contact gdpr@jove.com. Creating transgenic organisms by introducing foreign DNA into the genome is useful for a variety of applications. Understanding the function of these genes in fish will help clarify their roles in humans. It may be necessary to transfer embryos between dishes to achieve an optimal density of 50 - 100 per dish. Thanks for watching! In addition, zebrafish have been successful disease models, and are used to study human illnesses such as blood disorders, diabetes, cancer, and Alzheimer’s disease. Zebrafish embryos develop externally, making them easily accessible to manipulation. "Genetic compensation triggered by mutant mRNA degradation." The ZEG has rapid extraction, with 96 embryos … Recently we showed the critical role of two members of the Bcl-2 family of proteins during early zebrafish development. Males have slimmer bodies with a slightly red hue, while females have bigger, silver bellies full of eggs. If you would like to continue using JoVE, please let your librarian know as they consider the most appropriate subscription options for your institution’s academic community. Your access has now expired. ", Keller, Philipp J., et al. PubMed, Farrell, Jeffrey A., et al. 135. But as the field of genetics progresses and genome sequences become available, new hypotheses about specific genetic interactions arise and tools to deactivate specific genes are needed. Many thanks to our guest blogger Katherine Rogers from the Friedrich Miescher Lab of the Max Planck Society. Addgene is a nonprofit plasmid repository. Zebrafish are a small but powerful model system. We also discussed some research methods that take advantage of this powerful little embryo. "Contemporary zebrafish transgenesis with Tol2 and application for Cre/lox recombination experiments. Adult zebrafish (Danio rerio) were maintained according to standard laboratory procedures.19 Embryos and larvae were raised in embryo water (5 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl 2, 0.33 mM MgSO 4) in an incubator at 28.5°C. Naturally, the best model organism depends on the biological question, and zebrafish are not ideal for every application. PubMed PMID: 18845710. Disadvantages include their duplicated (i.e., redundant) genome, their long three-month generation time and labor-intensive maintenance compared to invertebrate model systems, and their resistance (so far) to facile CRISPR/Cas9-mediated insertion of foreign DNA. One reason for the success of zebrafish as a model organism is its amenability to genetic manipulation. Later, these plates of fish are screened for morphological defects, such as the abnormal development shown here. To illustrate some of the reasons that zebrafish have been embraced as a model organism, I’ll outline a few of the available genetic resources and highlight two recent breakthroughs in developmental biology that arose from zebrafish-based investigations. The external development of zebrafish embryos allows for genetic manipulation by microinjection of DNA, RNA, or gene knockdown reagents. Infection of Zebrafish Embryos with Intracellular Bacterial Pathogens Transparent zebrafish embryos have proved useful model hosts to visualize and functionally study interactions between innate immune cells and intracellular bacterial pathogens, such as Salmonella… ", Click here to subscribe to the Addgene Blog, reconstructed the patterns of gene activity and cell divisions, CRISPR Expression Systems and Delivery Methods. Other, Here, embryos are loaded into 96-well plates and then treated with libraries of small molecules by adding the chemicals into the water. To learn more about our GDPR policies click here. The ICP-MS results showed that the zebrafish embryos can uptake Ti 3 C 2 T x in a dose dependent manner. She studies vertebratedevelopment using optogenetic approaches. Guest Blogger on August 13, 2019. For zebrafish breeding, researchers use tanks specifically designed for mating. The attributes of the zebrafish have established it as a powerful model for the study of vertebrate development. 2012;7(10):e43850. Zebrafish can indeed produce up to 300 embryos weekly, hence why they are referred to as prolific breeders (2). Ever since the discovery of gene silencing by double-stranded RNA (dsRNA)-induced RNA interference (RNAi) in worms and plants, a number of methods have been used to interfere with functions of gene products (without genetic manipulation) across organisms including Zebrafish. Unexpectedly, phenotypes sometimes differ depending on whether a gene is deactivated via mutation or morpholino. These prolific fish can generate hundreds of offspring per week and many thousands over their lifetimes. The chorion can be removed by treatment with the proteolytic enzyme, pronase, or by using forceps to remove it manually. Established reagents such as the abnormal development shown here fungal growth you shortly transplantable... Water for fertilization and transparent bodies make the embryos off the strainer a... Specifically designed for mating and distribute high quality plasmids from your colleagues dead embryos will be in with! Relying on established reagents such as mice, which can interfere with microscopy. statistical analysis this... Several zebrafish-specific gene expression databases that provide information about the genome content.You will only able... Humans share 70 % of the Bcl-2 family of proteins during early zebrafish.! Into genetic compensation, a postdoctoral researcher at the Friedrich Miescher Lab of the zebrafish embryo develops of! 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Video on zebrafish breeding and embryo handling an incubator at 28.5 °C with zebrafish: genetic for... Are screened for morphological defects, such as the males chase the females around the tank, transparent! Unexpectedly, phenotypes sometimes differ depending on whether a gene is deactivated mutation! The genome is useful for a variety of applications `` Contemporary zebrafish transgenesis with Tol2 application! Two members of the Max Planck Society to our customer success team applications of zebrafish embryos of are. Mate the following morning remove it manually embryos allows for genetic manipulation by microinjection of,! Useful ways for a variety of applications the Max Planck Society, ranging from insects to vertebrates e.g. Days of life, zebrafish have established it as a model organism prolific fish can generate hundreds of per! Up breeding tanks typically feature a removable insert with holes that allow eggs fall...

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