Approaches for bacteriophage genome engineering

In recent years, bacteriophage research has been boosted by a rising interest in using phage therapy to treat antibiotic-resistant bacterial infections. In addition, there is a desire to use phages and their unique proteins for specific biocontrol applications and diagnostics. However, the ability to manipulate phage genomes to understand and...

One-step assembly and targeted integration of multigene constructs assisted by the I-SceI meganuclease in Saccharomyces cerevisiae

In vivo assembly of overlapping fragments by homologous recombination in Saccharomyces cerevisiae is a powerful method to engineer large DNA constructs. Whereas most in vivo assembly methods reported to date result in circular vectors, stable integrated constructs are often preferred for metabolic engineering as they are required for large-scale...

Single-molecule studies of the twisted, knotted, and broken genome

This thesis describes a series of single-molecule experiments aimed at understanding the physical properties of DNA itself and the proteins that interact with it. We developed and applied sensitive techniques that allowed us to directly probe the conformation and interactions of individual DNA molecules and proteins. Magnetic and optical...

Dynamic protein assemblies in homologous recombination with single DNA molecules

What happens when your DNA breaks? This thesis describes experimental work on the single-molecule level focusing on the interaction between DNA and DNA-repair proteins, in particular bacterial RecA and human Rad51, involved in homologous recombination. Homologous recombination and its central event of DNA strand exchange are essential processes...