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Figure 1 | BMC Biotechnology

Figure 1

From: Biotinylated-sortase self-cleavage purification (BISOP) method for cell-free produced proteins

Figure 1

Synthesis of srtA-fusion proteins using the wheat germ cell-free system. A. Schematic representation of the pEU-His-srtA-LPETG-Gene plasmids created using the Gateway system. B. Autoradiogram of SDS-PAGE of proteins synthesized using the cell-free system in the presence of [14C] Leu. Lane M, Protein MW standards labeled by using [14C]-containing felt pen. C. Autoradiogram of [14C] Leu incorporated GFP and SGK495 proteins synthesized by the wheat cell-free system in the presence of the Ca2+ chelating reagent BAPTA. The number represents concentration (mM) of BAPTA used in the protein synthesis reaction. Arrowheads denote the sizes of the full-length proteins. D. Rate of synthesis of the full-length protein and productivity of GFP (pink-colored bar and red-colored line) and SGK495 (purple-colored bar and blue-colored line) in the presence of different concentrations of BAPTA. E. Autoradiogram of [14C]-Leu incorporated proteins synthesized by the cell-free system in the presence of BAPTA. Asterisk denotes the sizes of the full-length proteins. F. Rate of synthesis of the full-length protein and productivity of proteins in the presence of different concentrations of BAPTA. Productivities of total synthesized and full-length proteins indicated as blue and red bars respectively. G. Purifications of proteins by the cell-free synthesis using the pEU-His-srtA-LPETG-Gene plasmid constructs. CBB-stained protein bands on the SDS-PAGE gel of the eluted (left panel) and resin-bound (right) target proteins are indicated using asterisk. Arrow represents the cleaved His-tagged srtA. Lane M (both panels): Protein MW standards.

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