Detection of Separase activity using a cleavage sensor in live mouse oocytes Detection of Separase activity using a cleavage sensor in live mouse oocytes
| Autor: | Nikalayevich, Elvira, Bouftas, Nora, Wassmann, Katja |
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| Přispěvatelé: | Institut de Biologie Paris Seine (IBPS), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: | |
| Zdroj: | Methods in Molecular Biology Methods in Molecular Biology, Humana Press/Springer Imprint, 2018 |
| ISSN: | 1064-3745 1940-6029 |
| Popis: | International audience; Separase proteolytically removes Cohesin complexes from sister chromatid arms, which is essential for chromosome segregation. Regulation of Separase activity is essential for proper cell cycle progression and correct chromosome segregation. Onset of Separase activity has not yet been observed in live oocytes. We describe here a method for detecting Separase activity in mouse oocytes in vivo. This method utilizes a previously described cleavage sensor made up of H2B-mCherry fused with Scc1(107-268 aa)-YFP. The cleavage sensor is loaded on the chromosomes through its H2B tag, and the signal from both mCherry and YFP is visible. Upon Separase activation the Scc1 fragment is cleaved and YFP dissociates from the chromosomes. The change in the ratio between mCherry and YFP fluorescence intensity is a readout of Separase activity. |
| Databáze: | OpenAIRE |
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