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Hox Genes pp 145-181 | Cite as

Transgenesis in Non-model Organisms: The Case of Parhyale

  • Zacharias Kontarakis
  • Anastasios Pavlopoulos
Part of the Methods in Molecular Biology book series (MIMB, volume 1196)

Abstract

One of the most striking manifestations of Hox gene activity is the morphological and functional diversity of arthropod body plans, segments, and associated appendages. Among arthropod models, the amphipod crustacean Parhyale hawaiensis satisfies a number of appealing biological and technical requirements to study the Hox control of tissue and organ morphogenesis. Parhyale embryos undergo direct development from fertilized eggs into miniature adults within 10 days and are amenable to all sorts of embryological and functional genetic manipulations. Furthermore, each embryo develops a series of specialized appendages along the anterior–posterior body axis, offering exceptional material to probe the genetic basis of appendage patterning, growth, and differentiation. Here, we describe the methodologies and techniques required for transgenesis-based gain-of-function studies of Hox genes in Parhyale embryos. First, we introduce a protocol for efficient microinjection of early-stage Parhyale embryos. Second, we describe the application of fast and reliable assays to test the activity of the Minos DNA transposon in embryos. Third, we present the use of Minos-based transgenesis vectors to generate stable and transient transgenic Parhyale. Finally, we describe the development and application of a conditional heat-inducible misexpression system to study the role of the Hox gene Ultrabithorax in Parhyale appendage specialization. Beyond providing a useful resource for Parhyalists, this chapter also aims to provide a road map for researchers working on other emerging model organisms. Acknowledging the time and effort that need to be invested in developing transgenic approaches in new species, it is all worth it considering the wide scope of experimentation that opens up once transgenesis is established.

Key words

Arthropods Crustaceans Parhyale hawaiensis Minos transgenesis Microinjections Heat-shock promoter Conditional gene misexpression Hox genes Ultrabithorax Appendage development 

Notes

Acknowledgements

We dedicate this chapter to the memory of Thanasis Loukeris, whose work paved the way for transgenic approaches in non-model organisms. We are grateful to Frederike Alwes for providing the drawings and photo shown in Fig. 2. Many protocols described in this chapter have been developed in close interaction with our Ph.D. supervisor and mentor Michalis Averof. Z.K. was supported by an EMBO long-term fellowship, and A.P. by a Marie Curie Intra-European fellowship and by the Howard Hughes Medical Institute.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Max Planck Institute for Heart and Lung ResearchBad NauheimGermany
  2. 2.Max Planck Institute of Molecular Cell Biology and GeneticsDresdenGermany
  3. 3.Howard Hughes Medical InstituteAshburnUSA

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