Improving CRISPR-Cas9 mediated genome integration in interspecific hybrid yeasts

Saccharomyces pastorianus is not a classical taxon, it is an interspecific hybrid resulting from the cross of Saccharomyces cerevisiae and Saccharomyces eubayanus. Exhibiting heterosis for phenotypic traits such as wort α-oligosaccharide consumption and fermentation at low temperature, it has been domesticated to become the main workhorse of...

Recognition of CRISPR mutational signatures using matrix factorization methods

Motivation: As one of the most common and life-threatening diseases in humans, cancer is a result of the accumulation of somatic mutations throughout the life cycle. Somatic mutation is a joint result of DNA lesion, which is a result of damage on DNA caused by mutagen, and the failure of DNA repair mechanisms. Therefore, understanding the...

Development of DNA diagnostics of neglected tropical diseases in resource-limited settings

The aim of this thesis was to develop a DNA-detection scheme for a point-of-care diagnostic test for Neglected Tropical Diseases (NTDs) for use within resource-limited settings. The scientific innovation is to develop an adaptable DNA-detection scheme, using CRISPR-dCas9 (catalytically inactive Cas9), that can detect the DNA of any pathogen in...

Genome editing in Kluyveromyces and Ogataea yeasts using a broad-host-range Cas9/gRNA co-expression plasmid

While CRISPR-Cas9-mediated genome editing has transformed yeast research, current plasmids and cassettes for Cas9 and guide-RNA expression are species specific. CRISPR tools that function in multiple yeast species could contribute to the intensifying research on non-conventional yeasts. A plasmid carrying a pangenomic origin of replication...

Detection of CRISPR-dCas9 on DNA with Solid-State Nanopores

Solid-state nanopores have emerged as promising platforms for biosensing including diagnostics for disease detection. Here we show nanopore experiments that detect CRISPR-dCas9, a sequence-specific RNA-guided protein system that specifically binds to a target DNA sequence. While CRISPR-Cas9 is acclaimed for its gene editing potential, the...

CRISPR-Cas9 mediated gene deletions in lager yeast Saccharomyces pastorianus

Background: The ease of use of CRISPR-Cas9 reprogramming, its high efficacy, and its multiplexing capabilities have brought this technology at the forefront of genome editing techniques. Saccharomyces pastorianus is an aneuploid interspecific hybrid of Saccharomyces cerevisiae and Saccharomyces eubayanus that has been domesticated for...

Genomic Determinants of Protein Abundance Variation in Colorectal Cancer Cells

Assessing the impact of genomic alterations on protein networks is fundamental in identifying the mechanisms that shape cancer heterogeneity. We have used isobaric labeling to characterize the proteomic landscapes of 50 colorectal cancer cell lines and to decipher the functional consequences of somatic genomic variants. The robust...

CRISPR/Cas9: A molecular Swiss army knife for simultaneous introduction of multiple genetic modifications in Saccharomyces cerevisiae

A variety of techniques for strain engineering in Saccharomyces cerevisiae have recently been developed. However, especially when multiple genetic manipulations are required, strain construction is still a time-consuming process. This study describes new CRISPR/Cas9-based approaches for easy, fast strain construction in yeast and explores their...