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Fig. 3 | Microbial Cell Factories

Fig. 3

From: Evaluation of factors influencing expression and extraction of recombinant bacteriophage endolysins in Escherichia coli

Fig. 3

Effect of inducer concentration on recombinant endolysins MatN and Cg solubility and yield. A, B E. coli C43(DE3) harboring pET-32a-MatN/Cg was induced at 20 °C by the addition of three different IPTG concentrations (0.1; 0.5 and 1 mM). Cells were harvested after 20 h of incubation with constant shaking and processed to obtain soluble (S) and insoluble (I) fractions. Equivalent protein amounts (10 µg) of each sample were loaded on 12% SDS-PAGE. Upon gel electrophoresis, gels were stained with Coomassie Brilliant Blue R-250 and unstained following standard protocols. Control (CTL) lanes correspond to cellular lysates of uninduced cultures. A sample of purified (His)6-MatN/Cg endolysin (57 kDa/57.7 kDa) was loaded on P lane. C, D Total amounts of recombinant endolysin (column height) and the ratio of soluble/insoluble protein (SP, gray dotted pattern; IP, diagonal lines pattern) were calculated after background normalization with ImageJ, open-source image processing software. Endolysin expression and solubility were analyzed in biological triplicates. Error bars represent SD of mean values

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