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Updated: Jul 31, 2025
Slice Preparation, Organotypic Tissue Culturing and Luciferase Recording of Clock Gene Activity in the Suprachiasmatic Nucleus
Published on: February 15, 2011
Detection of endogenous circadian rhythms of clock gene mRNA expression in mouse lung tissue using slice cultures
Ritsuko Matsumura1, Koichi Node2, Makoto Akashi1
1The Research Institute for Time Studies, Yamaguchi University, 1677-1 Yoshida, Yamaguchi, Yamaguchi 753-8511, Japan.
Abstract:
Detection of endogenous circadian rhythms in clock gene mRNA expression requires that mice be sacrificed at regular time intervals over one or more days. This protocol uses culture tissue slices obtained from a single mouse to collect time-course samples. We describe the procedure from preparation of lung slices to rhythmicity analysis of mRNA expression, including details to create handmade culture inserts. This protocol is useful for many mammalian biological clock researchers because it allows a decrease in animal sacrifice. For complete details on the use and execution of this protocol, please refer to Matsumura et al. (2022).1.
Insights
Researchers can now detect circadian rhythms in clock gene mRNA using tissue slices from a single mouse, reducing animal sacrifice. This method allows for time-course sample collection and rhythmicity analysis.
Area of Science:
- Molecular Biology
- Chronobiology
- Animal Research
Background:
- Detecting endogenous circadian rhythms in clock gene mRNA expression typically requires sacrificing multiple animals at various time points.
- This poses an ethical challenge and limits the scope of time-course studies.
Purpose of the Study:
- To present a novel protocol for detecting circadian rhythms in clock gene mRNA expression.
- To reduce the number of animal sacrifices required for circadian rhythm studies.
Main Methods:
- The protocol involves preparing cultured tissue slices from a single mouse to collect time-course samples.
- It details the procedure from lung slice preparation to mRNA expression rhythmicity analysis.
- Handmade culture inserts are described for facilitating the process.
Main Results:
- The described protocol enables the collection of time-course samples from a single mouse.
- It allows for the analysis of rhythmicity in clock gene mRNA expression.
Conclusions:
- This protocol offers a valuable alternative for mammalian biological clock researchers.
- It significantly decreases the number of animals that need to be sacrificed for circadian rhythm studies.
