条带转移(Band Shift)

 

 

 

Or gel mobility shift assay, gel shift assay, gel retardation, electrophoretic mobility shift assay (EMSA) 

• EMSA Using Oligos (Mike A. Dyer)
  Anneal two complementary oligos to make ds oligos as the probe for EMSA assay.
     
• Gel shift Assay (Hahn Lab)
    
• Band shifting (Promega)
  The gel shift assay is performed by incubating a purified protein, or a complex mixture of proteins (such as nuclear or cell extract preparations), with a 32P end-labeled DNA fragment containing the putative protein binding site. The reaction products are then analyzed on a nondenaturing polyacrylamide gel. The specificity of the DNA-binding protein for the putative binding site is established by competition experiments using DNA fragments or oligonucleotides containing a binding site for the protein of interest, or other unrelated DNA sequences.
    
• Bank Shift Assay for Kd Determinationi (Neri's Lab, Institute of Pharmaceutical Sciences ETHZ)
  The procedure is for the determination of affinity constants and kinetic dissociation constants by band-shift assay refers to an ideal antibody fragment (e.g., a scFv or an Fab fragment) binding to a well-behaved protein antigen (pure, of well-defined oligomeric state, migrating as a single band in non-denaturing gel electrophoresis. As mentioned in the introduction, either the antibody or the antigen has to be labeled in these assays.
     
• Fluorescent Band Shift Assay (Amersham)
  The band shift assay is a useful tool for identifying protein-DNA interactions and can be used to determine the affinity, abundance, binding constants, and binding specificity of DNA-binding proteins. The gel shift assay is performed by annealing two labeled oligonucleotides that contain the test binding sequence, then incubating the duplex with the binding protein. The mixture is then separated on a nondenaturing polyacrylamide gel. Duplexes that are bound by protein migrate more slowly than unbound duplexes and appear as bands that are shifted relative to the bands from the unbound duplexes.
  This protocol using fluorescein end-labeled oligonucleotides as probe for DNA-protein binding. Fluorescence imaging provides a rapid, sensitive, and quantitative alternative to radioactivity for performing band shift.