Abstract
Artificial cells made of molecular components and lipid membrane are emerging platforms to characterize living systems properties. Cell-free transcription-translation (TXTL) offers advantages for the bottom-up synthesis of cellular reactors. Yet, scaling up their design within well-defined geometries remains challenging. We present a microfluidic device hosting TXTL reactions of a reporter gene in thousands of microwells separated from an external buffer by a phospholipid membrane. In the presence of nutrients in the buffer, microreactors are stable beyond 24 h and yield a few mg/mL of proteins. Nutrients in the external solution feed the TXTL reaction at the picoliter scale via passive transport across the phospholipid membrane of each microfluidic well, despite the absence of pores. Replacing nutrients with an inert polymer and fatty acids at an isotonic concentration reduces microreactors efficiency, and a significant fraction yields no protein. This emphasizes the crucial role of the membrane for designing cell-free TXTL microreactors as efficient artificial cells.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1705-1712 |
| Number of pages | 8 |
| Journal | ACS Synthetic Biology |
| Volume | 8 |
| Issue number | 8 |
| DOIs | |
| State | Published - Aug 16 2019 |
Bibliographical note
Funding Information:We thank A. Libchaber and R. Sakamoto for discussion, and K. Tabata and H. Noji for microfabrication at the early stage of this study. This work was supported by Human Frontier Science Program Research Grant (RGP0037/2015), Grant-in-Aid for Scientific Research on Innovative Areas (JP16H00805 Synergy of Structure and Fluctuation, JP17H05234 Hadean Bioscience, and JP18H05427 Molecular Engines), and Grant-in-Aid for Scientific Research (B) JP17KT0025 from MEXT.
Publisher Copyright:
© 2019 American Chemical Society.
Keywords
- cell-free transcription-translation
- microfluidics
- passive trans-membrane transport