Recombinant Protein Expression and Purification

Recombinant plasmids were transformed into BL1 (DE3) cells for protein expression. They were then verified via Sanger DNA sequencing (Source Bioscience Ltd). The bacterial cells were grown as previously reported [28 (link)]. The GST-tagged recombinant protein was purified by affinity chromatography (columns) and quantified by the Bradford assay. Purified mutant proteins were analyzed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS–PAGE) gels to verify protein production.

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Akere A., Chen S.H., Liu X., Chen Y., Dantu S.C., Pandini A., Bhowmik D, & Haider S. (2020). Structure-based enzyme engineering improves donor-substrate recognition of Arabidopsis thaliana glycosyltransferases. Biochemical Journal, 477(15), 2791-2805.

Publication 2020

Sanger DNA sequencing

Affinity chromatography Assay Bacterial Cells Gels Mutant proteins Plasmids Protein Recombinant protein Sds page

Corresponding Organization : University College London

Other organizations : Oak Ridge National Laboratory, Sichuan Agricultural University, Brunel University of London

Top 5 similar protocols

Variable analysis

independent variables

Recombinant plasmids

dependent variables

Protein expression
Protein production

control variables

BL1 (DE3) cells
Growth conditions (as previously reported [28 (link)])
Affinity chromatography (columns) for protein purification
Bradford assay for protein quantification
SDS-PAGE analysis to verify protein production

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