Preparation of a Millimeter-Sized Supergiant Liposome That Allows for Efficient, Eukaryotic Cell-Free Translation in the Interior by Spontaneous Emulsion Transfer.

Abstract

We sought to prepare millimeter-sized supergiant 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 the 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.