| Solution Information | help | |
| Enzyme: | Agp1p | |
| inhibitor: | BDBM66026 | |
| substrate: | n/a | |
| Solution Type: | Aqueous | |
| pH at Preparation: | n/a | |
| Temp. Prep.: | n/a | |
| Comments: | The yeast cell-based multiplex assay is constructed using 5 strains from the Yeast-GFP Collection (Invitrogen, USA), representing 4 distinct branches of the TOR pathway: YAK Kinase, MSN4, GLN3 and RTG. Data were collected in single plex mode, meaning data from each individual strain were collected separately. Similar to the primary screening campaign, the assay is performed in a total volume of 10.1 microliters in 384-well microtiter plates. The strains are grown separately overnight in synthetic complete liquid media in a shaking incubator at 30 degrees C. Then each individual yeast strain was diluted into fresh media at 0.2 OD600. Aliquots of the yeast are transferred into 384-well microtiter plates and hit compounds found from primary screen are added in dose response concentrations ranging from 0.001 to 33 microM, final concentration. The cells are incubated for 3 hours at 30 degrees C with end-over-end rotation. Control wells contain the yeast strain treated for 3 hours with | |
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