Establishing a synthetic orthogonal replication system enables accelerated evolution in E. coli

被引：41

作者：

Tian, Rongzhen ^{[1
]}

Rehm, Fabian B. H. ^{[1
]}

Czernecki, Dariusz ^{[1
]}

Gu, Yangqi ^{[1
]}

Zurcher, Jerome F. ^{[1
]}

Liu, Kim C. ^{[1
]}

Chin, Jason W. ^{[1
]}

机构：

[1] MRC, Lab Mol Biol, Cambridge, England

来源：

SCIENCE | 2024年 / 383卷 / 6681期

基金：

英国医学研究理事会; 英国科研创新办公室;

关键词：

PRIMED DNA-REPLICATION; ESCHERICHIA-COLI; DIRECTED EVOLUTION; SOMATIC HYPERMUTATION;

D O I：

10.1126/science.adk1281

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

The evolution of new function in living organisms is slow and fundamentally limited by their critical mutation rate. Here, we established a stable orthogonal replication system in Escherichia coli. The orthogonal replicon can carry diverse cargos of at least 16.5 kilobases and is not copied by host polymerases but is selectively copied by an orthogonal DNA polymerase (O-DNAP), which does not copy the genome. We designed mutant O-DNAPs that selectively increase the mutation rate of the orthogonal replicon by two to four orders of magnitude. We demonstrate the utility of our system for accelerated continuous evolution by evolving a 150-fold increase in resistance to tigecycline in 12 days. And, starting from a GFP variant, we evolved a 1000-fold increase in cellular fluorescence in 5 days.

引用

页码：421 / 426

页数：6

共 40 条

[1] Development of potent in vivo mutagenesis plasmids with broad mutational spectra [J].