Cell-free proteins synthesis as a resource for generating plant proteins
The purpose of this project is to set up a cell-free protein synthesis (CFPS) resource based at Norwich BioScience Institutes in collaboration with University of Cambridge.
Cell-free protein synthesis (CFPS) has emerged as a powerful technique for on-demand, in vitro protein production which reduces labour and increases experimental throughput. However, cell-free systems can be laborious and expensive to set up and there is a shortage of publicly available data comparing different CFPS systems, particularly regarding plant proteins which can be difficult to express. This proposal will provide a resource for researchers to prototype their experimental plans without setting up the system themselves. We propose comparing an in-house generated E. coli S30 crude lysate system with a commercial wheat germ platform to quantify their ability to synthesize transcription factors and other plant proteins. This will provide data that can be used to build a simple model to predict which CFPS platform is best suited to a researcher’s needs. Additionally, two workshops (at Norwich and Cambridge) will communicate these results and provide additional expression data by crowdsourcing DNA assembly to workshop participants.
The purpose of this project is to set up a cell-free protein synthesis (CFPS) resource based at Norwich BioScience Institutes in collaboration with University of Cambridge. While there is growing interest in cell-free systems as a means for accelerating biological research, there is a higher barrier in terms of cost (£) and knowledge to setting up CFPS. Indeed, there is a shortage of publicly available information comparing the cost, yield, and flexibility of different CFPS systems. The limited data available often uses standardized non-plant reporter proteins such as green fluorescent protein (GFP) or luciferase which complicates comparison for plant biologists since plant proteins may require unique additives or an optimized folding environment for robust, soluble expression (Supplementary Figure 1 in proposal). This proposal aims to fill this knowledge gap by comparing a low-cost, in-house PANOx-SP E. coli S30 CFPS platform3 with a commercially available wheat germ4 CFPS kit from Promega.
Dr Quentin Dudley,
Postdoctoral Researcher, Engineering Biology Department, Earlham Institute, Norwich
Dr Susan Duncan,
Postdoctoral Researcher, Organisms and Ecosystems Department, Earlham Institute, Norwich
Mr Nicholas Larus-Stone,
Graduate Student, Department of Computer Science, University of Cambridge
This project is due to report in 2018.