Fig. 3

Growth curves in synthetic medium supplied with 2% galactose. a LmAcrIIA4-hosting circuits. The three working biosensors (A4-RGR, yo_A4-RGR, and yo_A4-pSNR52i) grew more slowly than byMM234, the negative control that expresses only yEGFP. By removing the gRNA from the circuit (dCas9-A4) or substituting it with a scrambled one with no match along the yeast genome (A4-RGR(scrambled) and yo_A4-RGR(scrambled)) a quick growth was re-established. b Biosensors based on LmAcrIIA2 confirmed that the RGR cassette is responsible for a slow growth when gRNAs targeting Tsynth8_pCYC1noTATA are expressed. In the presence of a scrambled gRNA (A2-RGR(scrambled)) as well as in the absence of any gRNA molecules (A2-dCas9) cell growth rate appeared even faster than that of the negative control. pSNR52 both on integrative (A2-pSNR52i) and multicopy (A2-pSNR52m) plasmid did not provoke drastic changes in cell growth either. c Both biosensors containing yo_StAcrIIA5 (yo_A5-RGR and yo_A5-pSNR52i) grew extremely slowly. However, the protein yo_StAcrIIA5 on its own has no negative effects on yeast cell growth as witnessed by the fast growth rate of the three control circuits (yo_A5, yo_A5-dCas9, and yo_A5-RGR(scrambled). Every average OD600 value was calculated on three replicates (single experiment)