Purification of Recombinant Proteins

Recombinant proteins were purified as previously described [18 (link)]. His6-tagged proteins were expressed in BL21 Escherichia coli cells. After isopropyl-β-d-thiogalactopyranoside (Sangon) induction, the cells were pelleted, lysed in PBS buffer, and incubated with Ni²⁺ TA beads (Sangon) to enrich the respective proteins, followed by elution with 500 mM imidazole (Sangon) dissolved in PBS buffer and then dialysed in PBS buffer supplemented with 20% glycerol before being aliquoted and preserved at −80°C.

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Zhao B., Yang Y., Cun B, & Chen P. (2021). AMBRA1 attenuates the proliferation of uveal melanoma cells. Open Medicine, 17(1), 1-14.

Publication 2021

isopropyl-β-d-thiogalactopyranoside Ni2+ TA beads imidazole

Buffer Cells Escherichia coli Glycerol Imidazole Proteins Recombinant proteins

Corresponding Organization : Kunming Medical University

Other organizations : Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences

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Variable analysis

independent variables

Expression of His6-tagged proteins in BL21 Escherichia coli cells
Induction with isopropyl-β-d-thiogalactopyranoside (Sangon)

dependent variables

Enrichment of respective proteins using Ni2+ TA beads (Sangon)
Elution with 500 mM imidazole (Sangon) dissolved in PBS buffer
Dialysis in PBS buffer supplemented with 20% glycerol

control variables

Lysis of cells in PBS buffer
Preservation of proteins at -80°C

controls

Positive control: Not specified
Negative control: Not specified

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