Practical Application of Microelectroporation into Developing Mouse Brain
One key approach toward understanding the genetic mechanisms underlying embryonic development involves the overexpression or misexpression of target genes in specific regions and at specific time points. The mouse gene-knockout system has been used extensively for loss-of-function studies due to the availability of a large number of mutant lines and the technical advantages of this system. In contrast, gain-of-function analyses have been performed through the production of knock-in and transgenic animals and with the use of various viruses (Cornetta 2006; Jakobsson et al., 2003; Hashimoto and Mikoshiba, 2004). However, it is not always possible to express or suppress genes in a spatially and temporally restricted manner, and the generation of genetically modified mice and recombinant viruses is time consuming and labor intensive. With the aim of solving these problems, many attempts have been made to apply the electroporation technique in research on developmental biology. Due to the accessibility of the avian embryo, it has been used as a classic model system for the study of developmental events in vertebrates. A novel technique for successful gene delivery into chick embryos has been established; this technique is known as in ovo electroporation and appears to be an excellent method, permitting quick and direct examination of the function of the delivered genes (Muramatsu et al., 1997; Itasaki et al., 1999; Momose et al., 1999; Nakamura et al., 2000; Yasuda et al., 2000). It seems that this technique can be adapted to the mouse embryo and would permit more rapid functional analysis of genes than is achieved by the generation of knockout or transgenic mouse lines. However, the inaccessibility of embryos in the mammalian uterus renders in utero manipulations targeting precise regions difficult or impossible at most stages of development. Efforts have been undertaken by various researchers to establish an in utero electroporation system, and there have been several reports of systems that enable successful gene delivery into mouse embryos, with time- and region-specific expression (Saito and Nakatsuji, 2001; Tabata and Nakajima, 2001; Borrell et al., 2005).
KeywordsUterine Horn Dorsal Cortex Develop Mouse Brain Ventral Telencephalon Zona Limitans Intrathalamica
Unable to display preview. Download preview PDF.
- Cornetta K, Pollok KE, Miller AD (2006) Retroviral vectors. Gene Transfer. Cold Spring Harbor Laboratory Press. Chapter 2. 3–21.Google Scholar
- Grove EA, Tole S, Limon J, Yip L, Ragsdale CW (1998). The hem of the embryonic cerebral cortex is defined by the expression of multiple Wnt genes and is compromised in Gli3-deficient mice. Development 125, 2315–25.Google Scholar
- Indra AK, Warot X, Brocard J, Bornert JM, Xiao JH, Chambon P, Metzger D (1999). Temporally-controlled site-specific mutagenesis in the basal layer of the epidermis: comparison of the recombinase activity of the tamoxifen-inducible Cre-ER(T) and Cre-ER(T2) recombinases. Nucleic Acids Res 27, 4324–7.CrossRefGoogle Scholar
- Maxwell IH, Maxwell F, Glode LM (1986). Regulated expression of a diphtheria toxin A-chain gene transfected into human cells: possible strategy for inducing cancer cell suicide. Cancer Res 46, 4660–4.Google Scholar
- Neumann E, Schaefer-Ridder M, Wang Y, Hofscneider PH (1982). Gene transfer into mouse lyoma cells by electroporation in high electric fields. EMBO J 1, 841–5.Google Scholar