Journal
ANALYTICAL BIOCHEMISTRY
Volume 439, Issue 2, Pages 80-87Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2013.04.019
Keywords
Clock gene; Secreted luciferase reporter; Circadian rhythm; Transgenic rats; In vivo recording
Funding
- Japan Society for the Promotion of Science (JSPS) KAKENHI [24390055, 20249010]
- Ministry of Education, Culture, Sports, Science, and Technology of the Japanese government
- Grants-in-Aid for Scientific Research [24390055] Funding Source: KAKEN
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A firefly luciferase reporter enabled us to monitor promoter activity in vivo as well as ex vivo; however, this requires a sufficient supply of the substrate luciferin and specific monitoring devices. To overcome these disadvantages, we developed transgenic rats carrying a secreted enzyme Cypridina luciferase (CLuc) reporter under the promoter of clock gene Per2 (Per2-CLuc). Per2-CLuc activity in serially sampled blood from freely moving rats exhibited robust circadian rhythms with a peak at early morning. The Per2-CLuc bioluminescence could be quantified even with approximately 100 pl of plasma. Plasma Per2-CLuc rhythms were phase reversed, and the level was reduced by restricting food access for 2 h during the light phase, suggesting that the plasma Per2-CLuc rhythms reflect the phase of peripheral clocks entrained to feeding cues as well as fuel metabolism. Fasting for 2 days did not alter the circadian Per2-CLuc rhythms in rats, suggesting that feeding per se did not affect the circadian Per2-CLuc rhythms. Tissue-specific Per2-CLuc rhythms were observed in culture medium of peripheral tissues. The Per2-CLuc reporter is a powerful tool to access gene expression in vivo as well as ex vivo with ordinary laboratory equipment. (C) 2013 Elsevier Inc. All rights reserved.
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