Preparation of Giant Unilamellar Vesicles as a Model to Study Viral Genome Packaging

Begin with a mixture of lipids that mimics the composition of a cell membrane, including a fluorescently tagged lipid.

Evaporate the solvent under vacuum to obtain a lipid film.

Add a sucrose solution to hydrate the dried film.

This organizes the lipid molecules into a bilayer.

Apply alternating electric fields, initiating bilayer swelling and the formation of giant unilamellar vesicles or GUVs, similar to the host cell membrane.

Store the GUVs in the sucrose solution to maintain their stability.

Next, incubate the fluorescently labeled viral proteins and RNA in buffered saline.

The viral protein recognizes and binds to the viral RNA, forming a viral protein–RNA complex.

Combine the vesicles with the viral protein–RNA complexes in a chambered coverglass.

The viral protein–RNA complexes interact with the vesicle surface, mimicking the early steps of viral particle assembly at the host cell membrane.

Under a confocal microscope, the GUVs appear as red rings with bright protein–RNA spots, confirming their membrane association.