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Protein membrane overlay assay: a protocol to test interaction between soluble and insoluble proteins in vitro.

J Vis Exp.. 2011-08;  (54):2961
Shoko Ueki, Benoît Lacroix, Vitaly Citovsky. Department of Biochemistry and Cell Biology, State University of New York.
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摘要

Validating interactions between different proteins is vital for investigation of their biological functions on the molecular level. There are several methods, both in vitro and in vivo, to evaluate protein binding, and at least two methods that complement the shortcomings of each other should be conducted to obtain reliable insights. For an in vivo assay, the bimolecular fluorescence complementation (BiFC) assay represents the most popular and least invasive approach that enables to detect protein-protein interaction within living cells, as well as identify the intracellular localization of the interacting proteins. In this assay, non-fluorescent N- and C-terminal halves of GFP or its variants are fused to test... More

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