Fig. 1 | Nature Communications

Fig. 1

From: Precise control of SCRaMbLE in synthetic haploid and diploid yeast

Fig. 1

Design of the “AND gate” switch for precise control of SCRaMbLE. a Design of the AND gate for SCRaMbLE and fitness assays in synV yeast and synIII yeast, respectively. Activation of the “AND gate” pCRE4 required the input of galactose and estradiol simultaneously. Fitness assays of synV yeast and synIII yeast containing pCRE1, pCRE4, and control vector (pRS413) separately. Strain growth was assessed at 30 °C on SC-His medium. Shown here are 10-fold serial dilutions after 2 d growth. b Transcriptional regulation and cellular localization regulation of Cre recombinase for tight control of SCRaMbLE. pCRE4 was activated by induction with galactose and estradiol. c Comparison of the growth of synV strains containing pCRE4 and pRS413 under four conditions as follows: (1) SC-His glucose medium without estradiol, (2) SC-His glucose medium with 1 μM estradiol, (3) SGal-His galactose medium without estradiol, and (4) SGal-His galactose medium with 1 μM estradiol. (Student’s t-test; NS, not significant; *P ≤ 0.05, **P ≤ 0.001). Error bars represent SD from three independent experiments. d Flow-cytometric assay expression level of Cre-EBD fusing GFPmut3b (488 nm excitation, 520 nm emission). Blue curves show fluorescence measurement of cells containing plasmids pRS413 incubated in SC-His glucose medium. Red curves show fluorescence measurement of cells containing plasmids pCRE5 incubated in SC-His glucose medium with 1 μM estradiol. Orange curves show fluorescence measurement of cells containing plasmids pCRE6 in SGal-His medium with 1 μM estradiol. e The “switch on” media for CRE5 and CRE6 were SC-His medium containing 1 µM estradiol and SGal-His medium containing 1 µM estradiol, respectively. The “initial state” and the “switch off” media were SC-His medium. The GFP fluorescence of the cells were imaged from this cell lawn using a fluorescence microscope (Olympus CX41). Scale bars, 10 µm

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