Single-molecule biophysics research group

University of California, Berkeley



 

Single-Molecule FRET

 

Single-molecule fluorescence resonance energy transfer (smFRET) is a powerful technique that enables us to observe interactions between proteins and nucleic acids, the structure of these complexes as well as their dynamic conformational changes at single molecule level. We label the samples site-specifically with a pair of fluorophores (called donor-acceptor pair) such that the emission spectrum of the donor overlaps with the excitation spectrum of the donor. Because of this overlap, when the donor is brought to its excited state with a laser, the excitation can be transferred to the acceptor, resulting in the emission of an acceptor photon. Since FRET efficiency is a function of the donor-acceptor separation, we can measure distances in the 2-10 nm range on a time scale ranging from milliseconds to several minutes.

smFRET trace

Figure 1. Anti-correlated donor and acceptor signals both attached to a DNA molecule and the calculated FRET efficiency. Dynamic conformational changes can be observed..

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