Abstract
Recent advances in molecular biology have led to the identification of several genes expressed at key stages of neural development. In order to evaluate gene function, scientists have typically used transgenic model systems or retroviral gene delivery systems. However, these methods are not always amenable to all experimental paradigms. Therefore, scientists have looked to develop alternative, non-viral methods of gene transfer that allow a wider range of approaches to study gene function, regulation, and expression (1). Recently, scientists have developed the technique of in vivo electroporation to misexpress genes in chick (2–13) and, to a limited extent, in mouse (2,14,15) embryos. Electroporation is an extremely effective method of targeting cells for gene misexpression, and can also be used to label a subset of cells within the neuroepithelium with a marker gene such as GFP.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Muramatsu, T., Mizutani, Y., Ohmori, Y., and Okumura, J. (1997) Comparison of three nonviral transfection methods for foreign gene expression in early chicken embryos in ovo. Biochem. Biophys. Res. Commun. 230, 376–380.
Itasaki, N., Bel-Vialar, S., and Krumlauf, R. (1999) ’Shocking’ developments in chick embryology: electroporation and in ovo gene expression. Nat. Cell. Biol. 1, E203–E207.
Araki, I. and Nakamura, H. (1999) Engrailed defines the position of dorsal di-mesencephalic boundary by repressing diencephalic fate. Development 126, 5127–5135.
Fujii, T., Funahashi, J., Matsuura, R., Izaki, T., Nakamura, H., Mikawa, T., and Tanaka, H. (2000) Axonal growth of the spinal cord interneurons expressing a homophilic adhesion molecule SC1 ectopically. Neurosci. Res. 38, 175–181.
Atkins, R. L., Wang, D., and Burke, R. D. (2000) Localized electroporation: a method for targeting expression of genes in avian embryos. Biotechniques 28, 94–96, 98,100.
DeLise, A. M. and Tuan, R. S. (2000) Electroporation-mediated DNA transfection of embryonic chick limb mesenchymal cells. Methods Mol. Biol. 137, 377–382.
Funahashi, J., Okafuji, T., Ohuchi, H., Noji, S., Tanaka, H., and Nakamura, H. (1999) Role of Pax-5 in the regulation of a mid-hindbrain organizer’s activity. Dev. Growth. Differ. 41, 59–72.
Katahira, T., Sato, T., Sugiyama, S., Okafuji, T., Araki, I., Funahashi, J., and Nakamura, H. (2000) Interaction between Otx2 and Gbx2 defines the organizing center for the optic tectum. Mech. Dev. 91, 43–52.
Koshiba-Takeuchi, K., Takeuchi, J. K., Matsumoto, K., Momose, T., Uno, K., Hoepker, V., Ogura, K., Takahashi, N., Nakamura, H., Yasuda, K., and Ogura, T. (2000) Tbx5 and the retinotectum projection. Science 287, 134–137.
Momose, T., Tonegawa, A., Takeuchi, J., Ogawa, H., Umesono, K., and Yasuda, K. (1999) Efficient targeting of gene expression in chick embryos by microelec-troporation. Dev. Growth. Differ. 41, 335–344.
Nakamura, H., Watanabe, Y., and Funahashi, J. (2000) Misexpression of genes in brain vesicles by in ovo electroporation. Dev. Growth. Differ. 42, 199–201.
Sakamoto, K., Nakamura, H., Takagi, M., Takeda, S., and Katsube, K. (1998) Ectopic expression of lunatic Fringe leads to downregulation of Serrate-1 in the developing chick neural tube; analysis using in ovo electroporation transfection technique. FEBS Lett. 426, 337–341.
Sugiyama, S., Funahashi, J., and Nakamura, H. (2000) Antagonizing activity of chick Grg4 against tectum-organizing activity. Dev. Biol. 221, 168–180.
Akamatsu, W., Okano, H. J., Osumi, N., Inoue, T., Nakamura, S., Sakakibara, S., Miura, M., Matsuo, N., Darnell, R. B., and Okano, H. (1999) Mammalian ELAV-like neuronal RNA-binding proteins HuB and HuC promote neuronal development in both the central and the peripheral nervous systems. Proc. Natl. Acad. Sci. USA 96, 9885–9890.
Yamazaki, Y., Yagi, T., Ozaki, T., and Imoto, K. (2000) In vivo gene transfer to mouse spermatogenic cells using green fluorescent protein as a marker. J. Exp. Zool. 286, 212–218.
Conrad, G. W., Bee, J. A., Roche, S. M., and Teillet, M. A. (1993) Fabrication of microscalpels by electrolysis of tungsten wire in a meniscus. J. Neurosci. Methods 50, 123–127.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2002 Humana Press Inc.
About this protocol
Cite this protocol
Van Raay, T.J., Stark, M.R. (2002). Cell Labeling and Gene Misexpression by Electroporation. In: Zigova, T., Sanberg, P.R., Sanchez-Ramos, J.R. (eds) Neural Stem Cells: Methods and Protocols. Methods in Molecular Biology™, vol 198. Humana Press, Totowa, NJ. https://doi.org/10.1385/1-59259-186-8:223
Download citation
DOI: https://doi.org/10.1385/1-59259-186-8:223
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-0-89603-964-3
Online ISBN: 978-1-59259-186-2
eBook Packages: Springer Protocols