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在大肠杆菌中由功能性TadA变体诱导的全基因组A → G和C → T突变

Hao Wang^1,2, Zhengxin Dong^1,2, Jingyi Shi^1,2

¹Laboratory of Synthetic Microbiology, School of Chemical Engineering & Technology, Tianjin University, Tianjin 300072, P. R. China.

ACS synthetic biology

|January 9, 2025

概括

研究人员开发了新的融合系统DnaG-TadA和DnaG-TadAD-UGI,以快速向细菌基因组引入多种突变. 这些系统显著加速了适应性进化,提高了大肠杆菌对抗生素和重金属的耐药性.

科学领域:

微生物学微生物学
遗传学遗传学是一个
合成生物学合成生物学

背景情况:

之前对细菌突变发生的研究主要使用细胞氨酸脱氨酶,将突变类型限制在C → T.
这种限制限制了通过随机突变发生可以实现的遗传多样性.

研究的目的:

开发新的融合表达系统,以增强大肠杆菌的随机突变发生.
在细菌基因组中引入更多种类的突变,特别是A → G和C → T.
将这些系统应用于加速适应性实验室进化.

主要方法:

DnaG (RNA原酶) 与腺因除氨酶TadA-8e (DnaG-TadA) 的融合表达,诱导A → G突变.
DnaG与双功能的TadA变体 (TadAD) 融合,并与 uracil glycosylase 抑制剂 (DnaG-TadAD-UGI) 共同表达,诱导 A → G 和 C → T 突变.
这些系统在适应性实验室进化实验中用于重金属和抗生素耐药性的应用.

主要成果:

DnaG-TadA系统将突变率提高了664倍,引入了A → G突变.
DnaG-TadAD-UGI系统将突变率提高了370倍,引入了A → G和C → T突变.
使用这些系统的适应性进化导致大肠杆菌显著增强了对Cd2+和卡纳米辛的耐药性,分别超过了传统方法的320%和266%.

结论:

新的DnaG-TadA和DnaG-TadAD-UGI融合系统为大肠杆菌的快速,多样化的随机突变产生提供了有效的策略.
这些系统加速了适应性实验室进化,从而显著改善了微生物耐药性表型.
开发的方法为各种 prokaryotic 物种的菌株改进提供了一个有希望的方法.

关键词:

埃舍里希亚大肠杆菌 (Escherichia coli) 是一个大肠杆菌.氨酸脱氨酸酶的使用.双基编辑双基编辑基因组的演化基因组的演化随机突变是一种随机突变.

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