DNA organization inside confined space

The genetic code of an organism is encoded on DNA that is present in every cell where it acts as the information carrier that provides the recipes for a cell to carry out its function. DNA is also a polymer, and a very special one indeed. Stretched out, it is many orders of magnitude larger than the cell to which it is confined. The very strongly compacted structure of the DNA influences its function. Supercoiling and nucleoid-associated proteins affect what genes are expressed or not. From in vivo studies, it became clear that the structure and dynamics of the genome are influenced by many parameters, such as geometric confinement, crowding, supercoiling and structural proteins.

We aim to deconvolve this complex parameter space by reconstituting the bacterial genome in artificial cells such as micro-fabricated chambers and droplets or liposomes, where we can control shape, concentrations, et cetera at will.

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Image: isolated bacterial nucleoid inside a 1.5 µm high chamber.

To achieve these aims, we isolate the bacterial nucleoid from the cell, remove all structural proteins, attach fluorescent labels along the contour as well as at predefined locations on the genome, and finally we encapsulate the isolated nucleoid in an artificial cell, such as a water-in-oil droplet, a micron-sized chamber or a liposome.

Using microfluidics the geometry of these artificial cells is manipulated and the effect of these spatial perturbations is observed on the dynamics of the encapsulated DNA. In addition to spatial perturbations, we also observe the influence of nucleoid associated proteins, by co-encapsulating purified versions of these along with the nucleoids in the artificial cells.