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Molecular, Biochemical, and Genetic Techniques

開催日 2014/9/11
時間 11:00 - 12:00
会場 Poster / Exhibition(Event Hall B)

Evaluation of a reporter rat line which conditionally expresses red fluorescent protein (tdTomato) under Cre-loxP system

  • P1-383
  • 五十嵐 敬幸 / Hiroyuki Igarashi:1,2 小泉 協 / Kyo Koizumi:2,3 金子 涼輔 / Ryosuke Kaneko:2,4 池田 啓子 / Keiko Ikeda:5 鬼丸 洋 / Hiroshi Onimaru:6 柳川 右千夫 / Yuchio Yanagawa:2,4 村松 慎一 / Shin-ichi Muramatsu:7 石塚 徹 / Toru Ishizuka:2,3 八尾 寛 / Hiromu Yawo:1,2,3 
  • 1:東北大院・医・神経細胞制御 / Dept Physiol and Pharmacol, Tohoku univ, Sendai. Japan 2:CREST, JST / CREST, JST 3:東北大院生命科学脳機能解析 / Dept Dev Biol and Neurosci, Tohoku univ, Sendai, Japan 4:群馬大院医遺伝発達行動 / Gunma Univ Grad Sch of Med, Maebashi, Japan 5:兵庫医科大学・生物学 / Hyogo College of Med, Nishinomiya, Japan 6:昭和大医第二生理 / Dept Physiol, Showa Univ Sch of Med, Tokyo, Japan 7:自治医科大 内科学講座 神経内科学部門 / Dept Med, Jichi Medical Univ, Tochigi, Japan 

The Cre/loxP recombination system is one of the conditional chromosomal mimics to investigate systemic function of targeted genes, and has been adopted to examine a function of specific gene. For in vivo experiments, rat offers potential advantages of larger body size and progressed ability to accomplish more complex behavioral task as compared to mouse. Here we evaluated a conditional reporter rat line which has red fluorescent protein (tdTomato) gene in the downstream of loxP-flanked STOP cassette.
Firstly we injected AAV-Cre into striatum, hippocampus and cerebellum of the reporter rats to test the conditional expression of tdTomato. Each Cre-immuno-positive cells sparsely located in the injection part and merged with tdTomato. Secondly, site-specific expression was evaluated by in utero electroporation of AcGFP-NCre plasmid. Cortical layer 2/3 neurons were visualized by tdTomato fluorescence in the newborn pup. Thirdly, we evaluated this reporter system using phox2b-Cre driver rat, which specifically expresses Cre in cells of several hindbrain regions involved in the autonomic nervous system including neurons that are responsible for respiratory rhythm generation. When the double transgenic rats with phox2b-Cre and floxed tdTomato were examined at embryonic day (E)12.5, tdTomato was expressed in the neurons of parafacial respiratory group (pFRG), epibranchial ganglia, the forming oculomotor/trochlear nuclei and autonomic ganglia.
It is suggested that the tdTomato was strongly expressed in neurons including Cre with high specificity. Our reporter rat would facilitate the neurophysiological studies and the connectomics of identified neurons which express Cre under a certain promoter.
All animal procedures were conducted in accordance with the guiding principles of Physiological Society of Japan and NIH.

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