Loss-of-function mutations and evolutionary stability dynamics in re-engineered T9002 circuits. (A) T9002 re-engineering involves changing the second double transcriptional terminator with varying degrees of homology and orientation to the first double transcriptional terminator. (B) Evolutionary stability dynamics of T9002 (solid black circles) and T9002 re-engineered circuits (various shapes and colors) under high input (+AHL) conditions. 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 re-engineered T9002 circuits. See Additional File 1, Supplementary Table S1 for mutation details.