Effects of Sugars on Giant Unilamellar Vesicle Preparation, Fusion, PCR in Liposomes, and Pore Formation
Kyoka Kajii, Ayu Shimomura, Mika Higashide, Masaya Oki, Gakushi Tsuji
Abstract
The water-in-oil emulsion transfer method was developed for preparing giant unilamellar vesicles (GUVs) and is useful for studying cellular functions under conditions that mimic cellular environments. A shortcoming of this method for encapsulating biochemical reactions is that it requires high sugar concentrations to enable the density effect to transverse the oil-water interface. In this study, we investigated the effects of sugars on GUV preparation and several biochemical reactions. We found that changing the sugar in the inner solution from sucrose to maltose or trehalose improved GUV formation. The fusion ratio of the freeze-thaw method was better in the traditional glucose-sucrose condition compared with the other examined conditions. For the inner biochemical reaction, we performed PCR in liposomes. The presence of maltose in the inner solution improved the stability of GUVs against damage caused by thermal cycles. Finally, fructose in the outer solution reduced leakage of the inner solution via pores on the membranes of GUVs. Our findings provide new insight for optimizing sugar conditions for preparing GUVs and inner GUV reactions. This could increase the utilization of GUVs as artificial cell compartment models.