A subscription to JoVE is required to view this content. Sign in or start your free trial.
Chapters
- 00:05Title
- 01:03Subculturing Cells and Physioxic Exposure
- 01:57Cell Lysis
- 05:06Cap-binding Assay
- 08:44Results: Analysis of Cap-binding Ability in Response to Oxygen of eIF4E and eIF4E2 in an m7GTP Affinity Column
- 09:50Conclusion
Here, we present human cell culture protocols to analyze translation initiation factors that bind the 5' cap of mRNA during physiological oxygen conditions. This method utilizes an Agarose-linked m7GTP cap analog and is suitable to investigate cap-binding factors and their interacting partners.
Tags
Cap-binding ProteinsPhysiological Oxygen ConditionsAgarose Linked Seven Methyl-guanosine Cap AnalogCap-binding FactorsProtein SynthesisOxygen Regulated FactorsCap Dependent TranslationHypoxiaCancer TherapyTumor ProgressionNormaxiaCell CultureLysis BufferProtease Inhibitor Cocktail4-benzenesulfonyl Fluoride HydroclorideTrypsin EDTA
