Shinmyo Yohei, Mito Taro, Matsushita Takashi, Sarashina Isao, Miyawaki Katsuyuki, Ohuchi Hideyo, Noji Sumihare
Department of Biological Science and Technology, Faculty of Engineering, The University of Tokushima, 2-1 Minami-Jyosanjima-cho, Tokushima City 770-8506, Japan.
Dev Growth Differ. 2004 Aug;46(4):343-9. doi: 10.1111/j.1440-169x.2004.00751.x.
Transgenic insects have been artificially produced to study functions of interesting developmental genes, using insect transposons such as piggyBac. In the case of the cricket, however, transgenic animals have not yet been successfully artificially produced. In the present study, we examined whether the piggyBac transposon functions as a tool for gene delivery in embryos of Gryllus bimaculatus. We used either a piggyBac helper plasmid or a helper RNA synthesized in vitro as a transposase source. An excision assay revealed that the helper RNA was more effective in early Gryllus eggs to transpose a marker gene of eGFP than the helper plasmid containing the piggyBac transposase gene driven by the G. bimaculatus actin3/4 promoter. Further, only when the helper RNA was used, somatic transformation of the embryo with the eGFP gene was observed. These results suggest that the piggyBac system with the helper RNA may be effective for making transgenic crickets.
利用诸如piggyBac等昆虫转座子，已人工培育出转基因昆虫以研究有趣的发育基因的功能。然而，就蟋蟀而言，尚未成功人工培育出转基因动物。在本研究中，我们检测了piggyBac转座子是否可作为一种在双斑蟋胚胎中进行基因传递的工具。我们使用piggyBac辅助质粒或体外合成的辅助RNA作为转座酶来源。一项切除试验表明，在双斑蟋早期卵中，辅助RNA在转座增强绿色荧光蛋白（eGFP）标记基因方面比含有由双斑蟋肌动蛋白3/4启动子驱动的piggyBac转座酶基因的辅助质粒更有效。此外，仅当使用辅助RNA时，才观察到胚胎被eGFP基因进行体细胞转化。这些结果表明，带有辅助RNA的piggyBac系统可能对培育转基因蟋蟀有效。
