Abstract
Cell-free protein synthesis (CFPS) is a platform technology that provides new opportunities for protein expression, metabolic engineering, therapeutic development, education, and more. The advantages of CFPS over in vivo protein expression include its open system, the elimination of reliance on living cells, and the ability to focus all system energy on production of the protein of interest. Over the last 60 years, the CFPS platform has grown and diversified greatly, and it continues to evolve today. Both new applications and new types of extracts based on a variety of organisms are current areas of development. However, new users interested in CFPS may find it challenging to implement a cell-free platform in their laboratory due to the technical and functional considerations involved in choosing and executing a platform that best suits their needs. Here we hope to reduce this barrier to implementing CFPS by clarifying the similarities and differences amongst cell-free platforms, highlighting the various applications that have been accomplished in each of them, and detailing the main methodological and instrumental requirement for their preparation. Additionally, this review will help to contextualize the landscape of work that has been done using CFPS and showcase the diversity of applications that it enables.
Highlights
Cell-free protein synthesis (CFPS) emerged about 60 years ago as a platform used by Nirenberg and Matthaei to decipher the genetic code and discover the link between mRNA and protein synthesis [1]
In an effort to assist the user in selecting the CFPS platform that is best suited to their experimental goals, this review provides an in-depth analysis of high adoption CFPS platforms in the scientific community, the applications that they enable, and methods to implement them
This review is aimed at helping new users of CFPS platforms determine which platform best suits their needs
Summary
Cell-free protein synthesis (CFPS) emerged about 60 years ago as a platform used by Nirenberg and Matthaei to decipher the genetic code and discover the link between mRNA and protein synthesis [1] Since this discovery, the CFPS platform has grown to enable a variety of applications, from functional genomics to large-scale antibody production [2,3]. The platforms have been categorized based on our understanding of their development and the degree to which they have been adopted by the field, as quantified by the number of peer-reviewed publications that utilize each platform (Figures 4B and 5B) This categorization allows new users to identify platforms that have been best established and to explore the applications that they enable. Low adoption platforms may be useful for niche applications, but have not been optimized thoroughly, or are
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