A microinjection protocol for the generation of transgenic killifish (Species: Nothobranchius furzeri)

• Published in: Developmental dynamics Vol. 241; no. 6; pp. 1133 - 1141
• Main Authors: Hartmann, Nils, Englert, Christoph
• Format: Journal Article
• Language: English
• Published: New York Wiley-Liss, Inc 01.06.2012; Wiley‐Liss, Inc; Wiley Subscription Services, Inc
• Subjects: aging; Animals; Animals, Genetically Modified - embryology; Animals, Genetically Modified - genetics; Embryonic Development - genetics; Freshwater; Gene Transfer Techniques; Genetic Vectors - genetics; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Killifishes; Microinjections - methods; Nothobranchius; short-lived teleost fish; tol2; transgenesis
• ISSN: 1058-8388; 1097-0177
• DOI: 10.1002/dvdy.23789

Background: A challenge in age research is the absence of short‐lived vertebrate model organisms. The turquoise killifish Nothobranchius furzeri has an exceptionally short lifespan of 4–10 months depending on the strain. Thus, it possesses the shortest known maximum lifespan of a vertebrate species that can be bred in captivity. Results: Here we show the successful introduction of DNA and RNA molecules into the one‐cell embryo of N. furzeri. For this purpose, we adapted existing microinjection protocols to inject through the remarkably thick and robust chorion of N. furzeri's eggs. The injected DNA transgene was integrated into the genome and transmitted to subsequent generations as indicated by the expression of the fluorophore enhanced green fluorescent protein (EGFP). Furthermore, we could confirm a special phase during embryonic development in which embryogenesis occurs within a re‐aggregated mass of previously dispersed cells as it has been described for other related cyprinodont fish species. Conclusions: The transgenesis protocol described here provides a basis for a variety of genetic manipulations including overexpression of genes and determining their effects on lifespan and longevity. The feasibility to perform transgenesis is an important step to establish N. furzeri as a new model in age research. Developmental Dynamics 241:1133–1141, 2012. © 2012 Wiley Periodicals, Inc. Key findings: A transgenesis protocol is presented for the short‐lived fish Nothobranchius furzeri. Successful germline transmission of the trangene is shown. N. furzeri possesses unique phases of embryonic development.