Journal
/
Biochimie
/
Toeprinting Analysis of Translation Initiation Complex Formation on Mammalian mRNAs
Journal JoVE
Biochimie
Un abonnement à JoVE est nécessaire pour voir ce contenu.  Connectez-vous ou commencez votre essai gratuit.
Journal JoVE Biochimie
Toeprinting Analysis of Translation Initiation Complex Formation on Mammalian mRNAs

Toeprinting Analysis of Translation Initiation Complex Formation on Mammalian mRNAs

DOI:
10.3791/57519-v

•

10:37 min

•

May 10, 2018

•

,
1Department of Chemistry and Biochemistry, Alberta RNA Research and Training Institute,University of Lethbridge, 2Department of Neuroscience and the Canadian Centre for Behavioral Neuroscience (CCBN),University of Lethbridge, 3Arnie Charbonneau Cancer Institute,University of Calgary

Chapitres

  • 00:04Titre
  • 00:34Preparation of mRNA
  • 02:59Toeprinting Assay
  • 06:32Sequencing Reactions and Gel Electrophoresis
  • 07:53Results: Importance of IRES Secondary Structure in the Formation of Translation Initiation Complexes on an Uncapped XIAP IRES RNA
  • 09:35Conclusion

Summary

Traduction automatique

Traduction automatique

Toeprinting aims to measure the ability of in vitro transcribed RNA to form translation initiation complexes with ribosomes under a variety of conditions. This protocol describes a method for toeprinting mammalian RNA and can be used to study both cap-dependent and IRES-driven translation.

Tags

Toeprinting AnalysisTranslation Initiation ComplexMammalian MRNAsIRESRNA PurificationRabbit Reticulocyte LysateRNase InhibitorATPGMP-PNPDTTSpermidine

Article

Embed

AJOUTER À LA PLAYLIST

Usage Statistics

Vidéos Connexes

Protein Complex Affinity Capture from Cryomilled Mammalian Cells

Preparation of Giant Vesicles Encapsulating Microspheres by Centrifugation of a Water-in-oil Emulsion

Quantitative Immunofluorescence to Measure Global Localized Translation

Genome-wide Quantification of Translation in Budding Yeast by Ribosome Profiling

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics

Complementation of Splicing Activity by a Galectin-3 – U1 snRNP Complex on Beads

Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex

Routine Collection of High-Resolution cryo-EM Datasets Using 200 KV Transmission Electron Microscope

Author Spotlight: Evaluating Biophysical Assays for Characterizing PROTACS Ternary Complexes

Author Spotlight: Characterization of Low-Affinity Protein Interactions in Solution Using MassFluidix Technology

Read Article