Figure 1 | Scientific Reports

Figure 1

From: Recombinant Fasciola hepatica fatty acid binding protein suppresses toll-like receptor stimulation in response to multiple bacterial ligands

Figure 1

Optimization of the Fh15 expression in E. coli TOP10. cDNA encoding Fh15 was cloned into the pGEX-4T-2 and expressed in E. coli bacteria as a fusion protein with glutathione S-transferase (GST) of Schistosoma japonicum at the amino end of protein. Small-scale protein expression using 4-ml of LB medium was induced for 3 h at 27 °C, 225 rpm. (A) Bacteria lysates induced at various concentrations of IPTG were analyzed by 15% SDS-PAGE Coomassie blue stained. (B) Unstained gel was transferred to nitrocellulose membrane and incubated with specific anti-GST antibody labeled with peroxidase. Arrow indicates maximal expression of the GST-tagged protein estimated to be ~41–43 kDa, observed at 0.02 mM IPTG. (C) Fusion protein was purified from a large-scale expression culture using a GSTrap FF 5 ml column in an AKTA FPLC System. Fusion protein is eluted using the elution buffer (EB): 10 mM Tris-HCl pH 8.0 containing 10 mM GSH. (D) The purification process was analyzed by 15% SDS-PAGE. Lane-1: GST-Fh15 fusion protein, lane-2: GST-tag and lane-3: Fh15. Figure 1A,B and D represent cropped images from the original and are being displayed in black & white. Original full-length gels and blots are shown in Supplementary Figure 1.

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