Fluorescence correlation spectroscopy as a detection tool of point mutation in genes

Fluorescence correlation spectroscopy (FCS) provides information about two important parameters in molecular biology. These are the average number of molecules in the detection volume, and the translational diffusion coefficient of the molecules. Although the properties of the translational diffusion reflect the molecular weight and shape, our work focuses on the analysis of the number of cleaved DNA fragments in solution. The cleavage of fluorescently labelled M13 DNA and pUC19 DNA was carried out with three kinds of restriction enzymes (HaeIII, HgaI and BsmAI), and the number of DNA fragments was monitored using FCS. The reciprocal value of the autocorrelation function at time τ = 0 is consistent with the number of fragments that are expected from restriction enzyme maps of the DNA. Our study indicates the capability of FCS as a tool for finding genetic differences in DNA sequences.