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Characterisation of a novel transgenic mouse for tissue- and cell-specific recording of circadian rhythms
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Garbutt, Lauren A (2022) Characterisation of a novel transgenic mouse for tissue- and cell-specific recording of circadian rhythms. PhD thesis, University of Warwick.
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Official URL: http://webcat.warwick.ac.uk/record=b3861719
Abstract
Reporters for the recording of circadian rhythms have provided significant advancements in our understanding of biological timing, from the intricacies of the molecular clock to complex behaviours such as sleep/wake and feeding,. Such reporters may include fluorescent tagging of circadian genes or inclusion of a bioluminescent construct which serves as a proxy for circadian gene dynamics. However, each cell in the mammalian system expresses clock genes and thus, the expression of the reporter is also ubiquitous. Therefore, a novel dual-reporter transgenic mouse was developed (DBPKI), utilising the CRE-Lox recombination system to allow the conditional induction of cell and tissue-specific bioluminescence via Luc2, driven by the clock controlled gene, Dbp. In the absence of CRE, Dbp expression is driving d2eGFP expression.
To characterise the functionality of this new reporter mouse as a tool for recording circadian rhythms, a series of in vivo, ex vivo and in vivo experiments were conducted using different models to ascertain the capabilities and limits of the knock-in construct. In doing so, it was found that the GFP aspect of the construct was not a reliable reporter for circadian timing, possibly due to the lack of a nuclear localisation signal and its destabilised nature. However, the luciferase construct was robustly rhythmic across all models and was specific enough to indicate tissue and indeed cell specificity, by observation of tanycyte circadian rhythms.
In a further step, the DBPKI reporter mouse was used to show that tanycytes, highly specific, nutrient responsive cells lining the third ventricle, do show cell autonomous circadian oscillations. Interestingly, the clock in tanycytes is sensitive to changes in nutrient levels suggesting an interaction between nutrient status and circadian timekeeping which may impact feeding behaviour.
Taken together, this DBPKI transgenic mouse is shown to be a valuable reporter for the circadian community, allowing us to interrogate tissue and cell-specific rhythms.
| Item Type: | Thesis (PhD) | ||||
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| Subjects: | Q Science > QH Natural history > QH426 Genetics Q Science > QP Physiology Q Science > QR Microbiology |
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| Library of Congress Subject Headings (LCSH): | Circadian rhythms, Circadian rhythms -- Molecular aspects, Transgenic mice, Biological rhythms, Bioluminescence, Luciferases | ||||
| Official Date: | April 2022 | ||||
| Dates: |
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| Institution: | University of Warwick | ||||
| Theses Department: | School of Life Sciences | ||||
| Thesis Type: | PhD | ||||
| Publication Status: | Unpublished | ||||
| Supervisor(s)/Advisor: | Dallmann, Robert ; Dale, Nicholas | ||||
| Sponsors: | Medical Research Council (Great Britain) ; University of Warwick. School of Life Sciences ; British Society for Neuroendocrinology | ||||
| Format of File: | |||||
| Extent: | 179 pages : illustrations | ||||
| Language: | eng |
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