Development of single-molecule tracking confocal microscope combined with force spectroscopy for gene-expression analysis

摘要

We have constructed a confocal fluorescence microscope combined with force measurements. Our method allows for simultaneous measurements of fluorescence anisotropy, energy transfer and correlation. The methodology and the sensitivity of the set-up using enhanced green fluorescent protein are described. We present results on (a) the detection of mRNA polymerization during in vitro transcription using fluorescence correlation spectroscopy and anisotropy, (b) detection of in vivo protein-DNA interactions using fluorescence anisotropy and (c) nanomanipulation of polytene chromosomes using the micropippete force sensor. Such a combined method allows for probing novel structure-function relationship underlying gene-expression.