TALENs-Assisted Multiplex Editing for Accelerated Genome Evolution To Improve Yeast Phenotypes

Genome
editing is an important tool for building novel genotypes
with a desired phenotype. However, the fundamental challenge is to
rapidly generate desired alterations on a genome-wide scale. Here,
we report TALENs (transcription activator-like effector nucleases)-assisted
multiplex editing (TAME), based on the interaction of designed TALENs
with the DNA sequences between the critical TATA and GC boxes, for
generating multiple targeted genomic modifications. Through iterative
cycles of TAME to induce abundant semirational indels coupled with efficient screening using a reporter, the targeted
fluorescent trait can be continuously and rapidly improved by accumulating
multiplex beneficial genetic modifications in the evolving yeast genome.
To further evaluate its efficiency, we also demonstrate the application
of TAME for significantly improving ethanol tolerance of yeast in
a short amount of time. Therefore, TAME is a broadly generalizable
platform for accelerated genome evolution to rapidly improve yeast
phenotypes.