Abstract
Over the last few years High-Throughput Protein Production (HTPP) has played a crucial role for functional proteomics. High-quality, high yield and fast recombinant protein production are critical for new HTPP technologies. Escherichia coli is usually the expression system of choice in protein production thanks to its fast growth, ease of handling and high yields of protein produced. Even though shake-flask cultures are widely used, there is an increasing need for easy to handle, lab scale, high throughput systems. In this article we described a novel minifermenter system suitable for HTPP. The Air-Well minifermenter system is made by a homogeneous air sparging device that includes an air diffusion system, and a stainless steel 96 needle plate integrated with a 96 deep well plate where cultures take place. This system provides aeration to achieve higher optical density growth compared to classical shaking growth without the decrease in pH value and bacterial viability. Moreover the yield of recombinant protein is up to 3-fold higher with a considerable improvement in the amount of full length proteins. High throughput production of hundreds of proteins in parallel can be obtained sparging air in a continuous and controlled manner. The system used is modular and can be easily modified and scaled up to meet the demands for HTPP.
Highlights
Over the last few years High-Throughput Protein Production (HTPP) has played a crucial role for functional proteomics
Overall features of the cultivation system Forced by a need to express and screen several hundred recombinant proteins in E. coli, we designed a system to allow growth of multiples of 96 cultures, at milliliterscale, in batch-mode with little instrumentation and labspace requirements
The key feature of the system is that simultaneous agitation/aeration of cultures is provided through a simple air sparging device (Figure 1A,B; AirWell minifermenter) powered by a low-pressure air supply
Summary
Over the last few years High-Throughput Protein Production (HTPP) has played a crucial role for functional proteomics. Even though shake-flask cultures are widely used, there is an increasing need for easy to handle, lab scale, high throughput systems. The increasing amount of data derived from genome scale studies has forced the implementation of protein expression pipelines that support high throughput (HT) screening and testing of cloned gene products [1,2] These pipelines combine batteries of cultivation devices eventually integrated with fluidic-handling and/or robotic apparatus for automated processes. Tubes or shake flasks have been the standard lab-scale cultivation devices for decades and have been used in industry and academia to grow a wide range of microorganisms as well as mammalian cells They are easy to operate and can be designed for automated monitoring of cultivation parameters such as pH and dissolved oxygen. The small specific mass transfer area of multiwell plates limits oxygen diffusion, leading to stress responses and medium acidification [6]
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