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Concept
Cell-free system
Summary
A cell-free system is an in vitro tool widely used to study biological reactions that happen within cells apart from a full cell system, thus reducing the complex interactions typically found when working in a whole cell. Subcellular fractions can be isolated by ultracentrifugation to provide molecular machinery that can be used in reactions in the absence of many of the other cellular components. Eukaryotic and prokaryotic cell internals have been used for creation of these simplified environments. These systems have enabled cell-free synthetic biology to emerge, providing control over what reaction is being examined, as well as its yield, and lessening the considerations otherwise invoked when working with more sensitive live cells.
Cell-free systems may be divided into two primary classifications: cell extract-based, which remove components from within a whole cell for external use, and purified enzyme-based, which use purified components of the molecules known to be involved in a given process. The cell extract-based type are susceptible to problems like quick degradation of components outside their host, as shown in a study by Kitaoka et al. where a cell-free translation system based on Escherichia coli (E. coli), of the cell extract-based type, had the mRNA template degrade very quickly and led to the halt of protein synthesis.
The methods of preparation vary between situations of both types of cell-free systems.
Nobel prize winner Eduard Buchner was arguably the first to present a cell-free system using yeast extracts, but since then alternative sources have been found. E. coli, wheat germ, and rabbit reticulocytes have all proven useful to create cell-free systems by extraction of their interior components. E. coli 30S extracts have been acquired, for example, by grinding the bacteria with alumina, followed by further cleaning. Similarly, wheat germ has been ground with acid-washed sand or powdered glass to open the cell membranes up. Rabbit reticulocytes have been lysed in a solution of MgCl_2 and had the extract filtered away from the membranes by centrifugation.
Official source
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