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. 2016 Sep 27;11(9):e0163511. doi: 10.1371/journal.pone.0163511

Fig 2. Expressing Gad proteins in Ty21a.

Fig 2

A. The three gad genes, gadA, gadB and gadC were cloned in tandem with their own ribosome binding sites under the control of an arabinose inducible promoter in a multi-copy plasmid pNW129 [37] to express these genes free from the typical complex regulatory network. B. The expression of Gad proteins was determined by Western blot with anti-Gad A/B antibody [35]. Gad A and Gad B are 98% similar at the protein level. The cloned GadA/B are very weakly expressed in the absence of arabinose induction, but Gad A/B are expressed in significant amounts as early as 2 hours after arabinose induction and their expression is increased over time. Gad A/B are not expressed in the negative control Ty21a containing the empty plasmid vector (Ty21a EV), but are expressed in overnight cultures of S. flexneri strain CFS100. A non-specific, antibody-binding band was used as the loading control; qualitative, rather than quantitative, levels of GadA/B expression were observed under different conditions.