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Figure 3 | BMC Biotechnology

Figure 3

From: Development of a Premature Stop Codon-detection method based on a bacterial two-hybrid system

Figure 3

Detection of inserts with PSCs through blue/white phenotypes in transformed BTH102 bacteria. Three different fragments, Wild Type (WT), Nonsense (NS), and Frameshift (FS), were obtained by mutagenic PCR from exon 11 of the human brca1 gene and inserted in pREAL. BTH102 transformed with pREAL were plated on LB-Xgal medium. All plates are shown on white and black backgrounds. In A, BTH102 were transformed with pREAL/WT. Colonies show the blue phenotype consistent with an active adenylate cyclase and a PSC-free DNA insert. In B, BTH102 were transformed with pREAL/FS, and in C, BTH102 were transformed with pREAL/NS. In the two latter cases, almost all colonies are white, indicating the absence of adenylate cyclase activity, and consistent with the presence of a PSC in the inserted DNA. In C we observed a few Lac+ colonies. A possible source of these false negative results is bacterial phenotype reversion due to endogenous mutations. Notice that this reversion rate does not disturb the diagnosis.

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