Expression and Purification of Tagged Protein in E.coli

E.coli has been found as a modal vector in which genes of different sources can be expressed. There have been many developments in the systems through which protein expression and purification can be achieved using E. coli as the cloning agent. Clontech HAT (Histidine Affinity Tag) is a protein expression and purification system whereby the vectors encode a novel polyhistidine epitope. This epitope allows the proteins that are expressed in bacteria to be purified at physiological pH at denaturing circumstances or conditions. This system presents two major advantages in that the proteins involved are soluble and hence they are no apparent aggregates within the inclusion bodies. The fact that the proteins can be washed out under neutral pH is also advantageous. (Scientifix, 2011)

Several purification strategies have also been devised. The clontech xTractor Buffer generally disrupts the bacterial cells to enhance protein purification. This system of protein purification has been optimized to enhance the extraction of poly-histidine tagged proteins. The extraction of protein using this method is simple. The cells are re-suspended in the buffer and mixed gently for about 10 minutes. The mixture of salts that results produces a lysate that has no visible precipitates. Proteins obtained through xTractor Buffer have a higher biological activity than cells that would be obtained through sonication. (Scientifix, 2011)

Other methods relate to protein expression and purification in E.coli and it is all a matter of choice depending on the characteristic of cell that one would want to achieve at the end of it. The choices are also dependent on the available time limit as well as financial resources at one’s disposal. However, the methods are eligible and result-oriented.

References

Scientifix, 2011, Protein Expression and Purification in E.coli, Web.

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StudyCorgi. (2022) 'Expression and Purification of Tagged Protein in E.coli'. 19 April.

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StudyCorgi. "Expression and Purification of Tagged Protein in E.coli." April 19, 2022. https://studycorgi.com/expression-and-purification-of-tagged-protein-in-e-coli/.

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StudyCorgi. 2022. "Expression and Purification of Tagged Protein in E.coli." April 19, 2022. https://studycorgi.com/expression-and-purification-of-tagged-protein-in-e-coli/.

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