Preparing Protein Producing Synthetic Cells using Cell Free Bacterial Extracts, Liposomes and Emulsion Transfer

Omer Adir; Noga Sharf-Pauker; Gal Chen; Maya Kaduri; Nitzan Krinsky; Janna Shainsky-Roitman; Jeny Shklover; Avi Schroeder

doi:10.3791/60829

Preparing Protein Producing Synthetic Cells using Cell Free Bacterial Extracts, Liposomes and Emu...

JoVE Journal
Bioengineering

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JoVE Journal Bioengineering

Preparing Protein Producing Synthetic Cells using Cell Free Bacterial Extracts, Liposomes and Emulsion Transfer

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09:37 min

April 27, 2020

DOI: 10.3791/60829-v

Omer Adir*^1,2, Noga Sharf-Pauker*^1,2, Gal Chen*^1,3, Maya Kaduri¹, Nitzan Krinsky^1,3, Janna Shainsky-Roitman¹, Jeny Shklover¹, Avi Schroeder¹

¹Laboratory for Targeted Drug Delivery and Personalized Medicine Technologies, Department of Chemical Engineering,Technion - Israel Institute of Technology, ²The Norman Seiden Multidisciplinary Program for Nanoscience and Nanotechnology,Technion - Israel Institute of Technology, ³The Interdisciplinary Program for Biotechnology,Technion - Israel Institute of Technology

This protocol describes the method, materials, equipment and steps for bottom-up preparation of RNA and protein producing synthetic cells. The inner aqueous compartment of the synthetic cells contained the S30 bacterial lysate encapsulated within a lipid bilayer (i.e., stable liposomes), using a water-in-oil emulsion transfer method.

Protein producing synthetic cells offer a versatile platform for studying the origins of life as well as synthetic cell-based therapies for advanced drug delivery. This method is simple and affordable for RNA and protein expression and can be applied for onsite therapeutic protein production directly inside the body. The systems can be used for treating a wide range of diseases from metabolic diseases and protein replacements to cancer and possibly diabetes.

This multi-step protocol has a defined stopping point. When performing it for the first time, it is recommended to divide the work over several days. The protocol for synthetic cell production starts with the preparation of the cell-free S30-T7 lysate.

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