Adaptive laboratory evolved BL21(DE3) by phloroglucinol challenge
Adaptive laboratory evolution was carried out using the GREACE method, in which mutated dnaQ gene (encoding proofreading factor of the DNA polymerase) was introduced into BL21(DE3) strain for in vivo continuous mutagenesis. After 52 rounds of selection, phloroglucinol concentration increased from 1 g/L to10 g/L, reaching its solubility, and the survival culture was spread onto LB agar plate to isolate single colonies. Then six colonies with the highest PG tolerance, which also showed improved phenol tolerance, were designated M01-06 and subjected to genome sequencing along with the parent BL21(DE3).