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Preparation of a mm-sized super-giant liposome that allows for efficient, eukaryotic cell-free translation in the interior by spontaneous emulsion transfer.

We sought to prepare mm-sized super-giant unilamellar vesicles (SGUVs) by spontaneous emulsion transfer for efficient, eukaryotic cell-free translation in the interior. Although the conventional protocols require that a considerably high concentration of sucrose be encapsulated into SGUVs for their efficient formation, such high amounts of sucrose severely inhibited cell-free translation based on wheat germ extract (WGE). We thus optimized the preparation conditions to permit SGUV formation at a much lower concentration of sucrose that has almost no effect on WGE translation. Under the optimized conditions, we successfully prepared WGE translation system-encapsulating SGUVs that allow for protein synthesis with a high efficiency comparable to that outside a liposome. The optimization also resulted in a high rate of successful SGUV formation (>90%) and a decent stability of the formed SGUVs (>60 min). These SGUVs are expected to serve as research tools in cell-free synthetic biology and as foundations for artificial cell-based biosensors.