Stable integration and conditional expression of electroporated transgenes in chicken embryos

Yuki Sato, Toshiharu Kasai, Shinichi Nakagawa, Koji Tanabe, Tadayoshi Watanabe, Koichi Kawakami, Yoshiko Takahashi

Research output: Contribution to journalArticle

174 Citations (Scopus)

Abstract

The in ovo electroporation in chicken embryos has widely been used as a powerful tool to study roles of genes during embryogenesis. However, the conventional electroporation technique fails to retain the expression of transgenes for more than several days because transgenes are not integrated into the genome. To overcome this shortcoming, we have developed a transposon-mediated gene transfer, a novel technique in chicken manipulations. It was previously reported that the transposon Tol2, originally found in medaka fish, facilitates an integration of a transgene into the genome when co-acting with Tol2 transposase. In this study, we co-electroporated a plasmid containing a CAGGS-EGFP cassette cloned in the Tol2 construct along with a transposase-encoding plasmid into early presomitic mesoderm or optic vesicles of chicken embryos. This resulted in persistent expression of EGFP at least until embryonic day 8 (E8) and E12 in somite-derived tissues and developing retina, respectively. The integration of the transgene was confirmed by genomic Southern blotting using chicken cultured cells. We further combined this transposon-mediated gene transfer with the tetracycline-dependent conditional expression system that we also developed recently. With this combined method, expression of a stably integrated transgene could be experimentally induced upon tetracycline administration at relatively late stages such as E6, where a variety of organogenesis are underway. Thus, the techniques proposed in this study provide a novel approach to study the mechanisms of late organogenesis, for which chickens are most suitable model animals.

Original languageEnglish
Pages (from-to)616-624
Number of pages9
JournalDevelopmental Biology
Volume305
Issue number2
DOIs
Publication statusPublished - May 15 2007
Externally publishedYes

Fingerprint

Transgenes
Chickens
Embryonic Structures
Transposases
Electroporation
Organogenesis
Tetracycline
Plasmids
Genome
Genes
Oryzias
Somites
Mesoderm
Southern Blotting
Embryonic Development
Retina
Cultured Cells
Fishes
Animal Models

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Developmental Biology
  • Cell Biology

Cite this

Sato, Y., Kasai, T., Nakagawa, S., Tanabe, K., Watanabe, T., Kawakami, K., & Takahashi, Y. (2007). Stable integration and conditional expression of electroporated transgenes in chicken embryos. Developmental Biology, 305(2), 616-624. https://doi.org/10.1016/j.ydbio.2007.01.043

Stable integration and conditional expression of electroporated transgenes in chicken embryos. / Sato, Yuki; Kasai, Toshiharu; Nakagawa, Shinichi; Tanabe, Koji; Watanabe, Tadayoshi; Kawakami, Koichi; Takahashi, Yoshiko.

In: Developmental Biology, Vol. 305, No. 2, 15.05.2007, p. 616-624.

Research output: Contribution to journalArticle

Sato, Y, Kasai, T, Nakagawa, S, Tanabe, K, Watanabe, T, Kawakami, K & Takahashi, Y 2007, 'Stable integration and conditional expression of electroporated transgenes in chicken embryos', Developmental Biology, vol. 305, no. 2, pp. 616-624. https://doi.org/10.1016/j.ydbio.2007.01.043
Sato, Yuki ; Kasai, Toshiharu ; Nakagawa, Shinichi ; Tanabe, Koji ; Watanabe, Tadayoshi ; Kawakami, Koichi ; Takahashi, Yoshiko. / Stable integration and conditional expression of electroporated transgenes in chicken embryos. In: Developmental Biology. 2007 ; Vol. 305, No. 2. pp. 616-624.
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