To gain a better understanding of material properties of chromatin and successfully link them to chromatin organization and functions such as transcription, we develop novel methods to actively manipulate a genomic locus inside the nucleus of a living human cell. By targeting iron-containing nanoparticles to a specific genomic locus and applying a controlled magnetic field, we were able to physically move chromatin through nuclear space for the first time. Exertion of near-picoNewton forces led to displacements over microns within minutes. We observe partially reversible stretching of chromatin highlighting its’ viscoelastic nature. We could accurately recapitulate the observed behavior with a Rouse model that included only a weak obstructive effect of the surrounding chromatin and nucleoplasmic material. This challenges the view that interphase chromatin is a gel-like material.
Water-soluble quantum dots (qdots) have been introduced as bright fluorophores into life sciences research. Although various photophysical pathologies of qdots have been found, how their biological applications will be affected --- particularly in the native biological environment --- has not been evaluated. By fluorescence coincidence analysis and fluorescence cross-correlation spectroscopy, we studied the dark fraction of free-diffusing qdots in aqueous solution. We were able to detect individual qdots and found significant heterogeneity --- well-distinguished dark qdots and bright qdots. We estimated the bright fraction of Qdot525-Streptavidin to be about 55%. Blinking events were also noticed in fluorescence coincidence analysis, with “on/off” timescale from submilliseconds to tens of milliseconds.
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