Screening target microorganisms from a mutated recombinant
library
plays a crucial role in advancing synthetic biology and metabolic
engineering. However, conventional screening tools have several limitations
regarding throughput, cost, and labor. Here, we used the fluid array
platform to conduct high-throughput screening (HTS) that identified Escherichia coli ‘TesA thioesterase mutants
producing elevated yields of free fatty acids (FFAs) from a large
(106) mutant library. A growth-based screening method using
a TetA-RFP fusion sensing mechanism and a reporter-based screening
method using high-level FFA producing mutants were employed to identify
these mutants via HTS. The platform was able to cover >95% of the
mutation library, and it screened target cells from many arrays of
the fluid array platform so that a post-analysis could be conducted
by gas chromatography. The ‘TesA mutation of each isolated
mutant showing improved FFA production in E. coli was characterized, and its enhanced FFA production capability was
confirmed.