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Chapters
- 00:05Title
- 01:30Transfection
- 02:20Cell Lysis
- 03:30Binding to Rabbit IgG-agarose
- 04:53TEV Protease Cleavage
- 05:48Binding to Immobilized Streptavidin Beads
- 07:50Biotin Elution of the Streptavidin Binging Peptide and Bait Protein
Tandem affinitetsoprensning er en robust fremgangsmåde til identifikation af protein-bindende partnere. Som proof of concept, blev denne metode anvendt på velkarakteriserede translationsstart faktor eIF4E til co-bundfald værtscellesystemerne faktorer involveret i oversættelse initiering. Denne fremgangsmåde kan let tilpasses til enhver cellulært eller viralt protein.
Tags
Protein Interacting PartnersTandem Affinity PurificationIdentification Of Interacting ProteinsYeast Two Hybrid SystemPull Down AssaysImmunoprecipitationMass Spectrometry IdentificationDouble-affinity Tag Mediated PurificationTAPEukaryotic Translation Initiation Factor EIF4ECellular Translation Factor EIF4EEIF4F ComplexCap-dependent Translation InitiationProtein G UnitsStreptavidin Binding PeptideTobacco Etch Virus (TEV) Protease Cleavage Sequence
