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Fig. 5 | BMC Plant Biology

Fig. 5

From: Determinants of substrate specificity in a catalytically diverse family of acyl-ACP thioesterases from plants

Fig. 5

Recombinant ALT-like thioesterases are expressed successfully in K27(DE3) E. coli. Left: Immunoblots of crude cell lysates from K27(DE3) E. coli strains expressing wild-type or mutant ALT thioesterases show a prominent band at ~ 20 kDa, which is not present in samples from a strain harbouring an empty pET28a vector. 10 μg of total protein was loaded into each lane. Expressed ALTs carried an N-terminal T7-tag, and ALT protein bands were detected by probing the membrane with an anti-T7 mouse monoclonal primary antibody and an anti-mouse horseradish peroxidase-conjugated secondary antibody. All membranes were imaged at 59 s exposure using a BioRad ChemiDoc XRS+ system with ImageLab v6.0.1 software. Images taken from different membranes are separated. Right: Membranes used for immunodetection of ALT proteins stained with Ponceau S following transfer from Tris-Tricine SDS-PAGE gels. All Ponceau-stained membranes were imaged under white light, with an exposure time of 1/30 s. Full-length membranes are displayed in Fig. S4

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