Systematic genome engineering approaches to investigate mutational effects and evolutionary processes

Nyerges Ákos József
Systematic genome engineering approaches to investigate mutational effects and evolutionary processes.
[Thesis] (Unpublished)

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Abstract in foreign language

To address the shortcomings of currently available genome editing and in vivo directed evolution techniques, we have developed a plasmid-based method for broad-host-range genome engineering (pORTMAGE), and based on pORTMAGE, a method for in vivo directed evolution. This new method, termed DIvERGE (directed evolution with random genomic mutations) allows the systematic multiplex mutagenesis of long genomic segments. DIvERGE has numerous advantages over the alternative techniques, including (I) the possibility to target multiple, user-defined genomic regions; (II) it has a broad and controllable mutagenesis spectrum for each nucleotide position; (III) it allows of up to a million-fold increase in mutation rate at the target sequence; (IV) it enables multiple rounds of mutagenesis and selection in a fast and continuous manner; (V) it is applicable to a wide range of enterobacterial species without the need for prior genomic modification(s); (VI) it avoids off-target mutagenesis, and (VII) it is also cost-effective as it relies on soft-randomized oligos which can easily be manufactured at a modest cost. In summary, DIvERGE offers a versatile solution for high-precision directed evolution at multiple loci in their native genomic context. Due to these favorable characteristics, DIvERGE is especially well-suited to study bacterial evolution leading to antibiotic resistance.

Item Type: Thesis (PhD)
Creators: Nyerges Ákos József
Hungarian title label: Rendszerszintű genommérnöki vizsgálatok az evolúciós folyamatok tanulmányozására
Divisions: Doctoral School of Biology
Discipline label: Natural Sciences > Biology
Defence date label: 2019. March 18.
Uncontrolled Keywords: Synthetic biology, genome engineering, antibiotic resistance, directed evolution
Item ID: 10051
MTMT id: 30435535
Date Deposited: 2019. Jan. 09. 19:28
Last Modified: 2020. Jul. 06. 10:43
Depository no.: B 6501
Defence/Citable status: Defended.

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