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. 2014 Sep 16;111(39):14241–14246. doi: 10.1073/pnas.1411570111

Fig. 3.

Fig. 3.

The toxT 5′ UTR fourU RNA thermometer facilitates temperature-dependent translation of ToxT. (A) Schematic representation of the 5′-toxT UTR-toxT-FLAG transcript of pKEK1733. E. coli DH5α (E. coli), V. cholerae O1 ΔtoxT strains KKV2426 (Vc El Tor) and KKV2288 (Vc Classical) carrying plasmid pKEK1733 were grown at the indicated temperatures in the presence of 0.1% arabinose. (B) Schematic representation of the 5′-araBAD-UTR-toxT-FLAG transcript of pKEK1789. E. coli DH5α (E. coli), V. cholerae O1 strain C6706 (Vc El Tor), and V. cholerae O1 strain O395 (Vc Classical) carrying plasmid pKEK1789 (5′-araBAD-UTR) were grown at the indicated temperatures in the presence of 0.1% arabinose. Western immunoblot using anti-FLAG monoclonal antibody was performed in A and B with whole-cell lysates (OD600 = 0.6). (C) Mutations in the fourU thermometer disrupt temperature-dependent translation of ToxT. E. coli DH5α carrying plasmids pKEK1733 (5′-toxT UTR), pKEK1734 (5′-U5C toxT UTR), pKEK1735 (5′-U7C toxT UTR), or pKEK1736 (5′-U5C U7C toxT UTR) were grown at 37 °C in the presence of 0.1% arabinose. Western immunoblot using anti-FLAG monoclonal antibody was performed with whole-cell lysates (OD600 = 0.6).