Loss-of-function mutations and evolutionary stability dynamics in two lacI -regulated circuits under constitutive vs. inducible expression. (A) Regulation of inducible R0011 + E0240 and R0010 + E0240 circuits. LacI represses transcription of GFP and IPTG de-represses the circuit to allow for GFP expression. (B) Top panel shows the evolutionary stability dynamics of constitutive R0011 + E0240 (open black circles) and inducible R0011 + E0240 (solid black circles). Bottom panel shows the evolutionary stability dynamics of constitutive R0010 + E0240 (open red circles) and inducible R0010 + E0240 (solid red circles). Error bars represent one standard deviation from the mean of nine independently evolved populations. (C) Types of mutations in nine independently evolved populations. For nine independently evolved populations, colored boxes correspond to the mutation legend below the table. The most common mutation for a particular type of mutation is labeled with "1" in the boxes above and less common mutations are labeled with increasing numbers. (D) Most common loss-of-function mutations that inactivated the R0011 + E0240 and R0010 + E0240 inducible and constitutive circuits. See Additional File 1, Supplementary Table S1 for mutation details.