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Establishment of cells to monitor Microprocessor through fusion genes of microRNA and GFP |
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| Authors: | Tsutsui Motomu Hasegawa Hitoki Adachi Koichi Miyata Maiko Huang Peng Ishiguro Naoki Hamaguchi Michinari Iwamoto Takashi |
| Affiliation: | a Department of Orthopedic Surgery, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Sowa-ku, Nagoya, Aichi 466-8550, Japan b The Center for Education in Laboratory Animal Research, Chubu University, 1200 Matsumoto-cho, Kasugai, Aichi 487-8501, Japan c Radioisotope Research Center Medical Division, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Sowa-ku, Nagoya, Aichi 466-8550, Japan d Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, 1200 Matsumoto-cho, Kasugai, Aichi 487-8501, Japan e Department of Biochemistry, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Sowa-ku, Nagoya, Aichi 466-8550, Japan f Division of Cancer Biology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Sowa-ku, Nagoya, Aichi 466-8550, Japan |
| Abstract: | Microprocessor, the complex of Drosha and DGCR8, promotes the processing of primary microRNA to precursor microRNA, which is a crucial step for microRNA maturation. So far, no convenient assay systems have been developed for observing this step in vivo.Here we report the establishment of highly sensitive cellular systems where we can visually monitor the function of Microprocessor. During a series of screening of transfectants with fusion genes of the EGFP cDNA and primary microRNA genes, we have obtained certain cell lines where introduction of siRNA against DGCR8 or Drosha strikingly augments GFP signals. In contrast, these cells have not responded to Dicer siRNA; thus they have a unique character that GFP signals should be negatively and specifically correlated to the action of Microprocessor among biogenesis of microRNA.These cell lines can be useful tools for real-time analysis of Microprocessor action in vivo and identifying its novel modulators. |
| Keywords: | microRNA Drosha DGCR8 Dicer Microprocessor RNA interference Green fluorescent protein |
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