RT Journal Article SR Electronic T1 Configurable Compartmentation Enables In Vitro Reconstitution of Sustained Synthetic Biology Systems JF bioRxiv FD Cold Spring Harbor Laboratory SP 2022.03.19.484972 DO 10.1101/2022.03.19.484972 A1 Luyao Li A1 Rong Zhang A1 Xintong Tian A1 Ting Li A1 Bingchun Pu A1 Conghui Ma A1 Fang Ba A1 Chenwei Xiong A1 Yunfeng Shi A1 Jian Li A1 Jay Keasling A1 Jingwei Zhang A1 Yifan Liu YR 2022 UL http://biorxiv.org/content/early/2022/05/11/2022.03.19.484972.abstract AB The compartmentalized and communicative nature of biological cells contributes to the complexity and endurance of living organisms. Current in vitro compartmentalization systems such as droplet emulsions reproduce the compartmentalization property of cells yet fail to recapture the configurability of cellular communication with the environment. To mimic biological cells a step further and expand the capabilities of in vitro compartmentalization, we present here a general strategy that inherits the passive transport phenomenon of biology. The strategy incorporates layered, micrometer-sized, hydrogel-based compartments featuring configurability in composition, functionality, and selective permeability of biomolecules. We demonstrated the unique advantage of our strategy in two scenarios of synthetic biology. First, a compartmentalized cell-free protein synthesis system was reconstituted that could support multiple rounds of reactions. Second, we constructed living bacteria-based biosensors in the hydrogel compartments, which could achieve long-lasting functioning with markedly enhanced fitness in complex environments. Looking forward, our strategy should be widely applicable for constructing complex, robust, and sustained in vitro synthetic molecular and cellular systems, paving the way for their practical applications.Competing Interest StatementA patent of methods described in this paper is under application.