Current Protocols in Molecular Biology
Featured Protocol

Figure 20.1.5 Repression assay for DNA binding. (A) The plasmid JK101 contains the upstream activating sequence (UAS) from the GAL1 gene followed by LexA operators upstream of the lacZ coding sequence. Thus, yeast containing pJK101 will have significant b-galactosidase activity when grown on medium in which galactose is the sole carbon source because of binding of endogenous yeast GAL4 to the GALUAS (B). LexA-fused proteins (P1-LexA) that are made, enter the nucleus, and bind the LexA operator sequences (ops) will block activation from the GALUAS, repressing b-galactosidase activity (+) 3- to 5-fold. On glucose/Xgal medium, yeast containing pJK101 should be white because GALUAS transcription is repressed.