Room-temperature Storage of Lyophilized Engineered Bacteria using Tardigrade Intrinsically Disordered Proteins

Abstract

Tardigrades, which live in transiently wet environments such as moss, are well-known for their extreme resistance to desiccation. Tardigrade intrinsically disordered proteins (TDPs) have been reported to also protect bacteria and yeast under desiccation. In this study, we utilized lyophilization to achieve room-temperature storage of engineered bacteria. By using TDPs, engineered bacteria are protected under lyophilization and their original functions are preserved. This study shows that TDPs can be expressed in the Escherichia coli (E. coli) BL21 and DH5α, and bacteria treated with Cytosolic-abundant heat soluble protein (CAHS) 106094 displayed the highest survival rate after lyophilization. Moreover, this study shows that the co-expression of TDPs can improve the preservation of bacteria and maintain high survival rates after prolonged room temperature storage. Additionally, the TDPs can be expressed using different vectors, which means that they can be used in different types of engineered bacteria. This study offers a new storage method that not only improves the storage of biological material for industrial and daily usage, but also for future iGEM (International Genetically Engineered Machine Competition) teams to store and use their engineered bacteria in different applications.