We develop CRISPR-based technologies for high-throughput genome and transcriptome perturbation in microbes, and we apply these technologies in the model eukaryote Saccharomyces cerevisiae to study complex eukaryotic genetics.
Through advances in sequencing technologies we are now able to generate detailed maps of sequence variation across entire genomes. However, our knowledge on the functional consequences of the majority of these variants is lagging behind. Methods for precise and accurate manipulation of genomes enable assessing the function of genes and gene variants directly, analyzing their contribution to phenotypes of interest, and engineering cells with desired characteristics. RNA guided programmable DNA nucleases of CRISPR systems, in particular the type II CRISPR/Cas9 system from Streptococcus pyogenes, have been widely adopted for efficient and precise genome manipulation. The Cas9 nuclease can be directed to cut any genomic location using a short guide RNA (gRNA) with homology to the target region if a protospacer-adjacent motif (PAM) is next to the target sequence. Following gRNA-directed, Cas9-mediated cutting of the genome, donor DNA templates encoding desired mutations flanked by homologies to the target site are used in homologous recombination (HR)-mediated repair of the cut. Array-based oligonucleotide synthesis allows fast and affordable creation of guide-donor libraries directed against thousands of targets.
In the lab we develop highly multiplexed CRISPR/Cas based technologies for precision genome engineering in microbes. Our technologies allow us to engineer thousands of variants in parallel in a single test tube. We apply our tools to study how sequence variation shapes phenotypic complexity and to screen variants for advantageous properties in industrial applications. We also develop scalable and cost-efficient technologies for genomic and functional characterization of our edited strains. We mainly work with the yeast S. cerevisiae, which is a model for many basic processes in humans and other biological systems and an industrially important microbe.
High-throughput CRISPR-based precision genome editing
A major focus in the lab is the development of CRISPR technologies for massively parallel precision genome editing and direct functional screening of sequence variation in microbes. This builds on our multiplexed variant engineering and barcode-based phenotyping platform MAGESTIC (Multiplexed Accurate Genome Editing with Short, Trackable, Integrated Cellular barcodes), which uses pools of array-synthesized oligos encoding a gRNA and a donor DNA to introduce a designed variant by HR. Importantly, variant strains are tagged by DNA barcodes integrated at a dedicated genomic locus, allowing to efficiently track mutations in cell populations during functional screens, and read out variant identities via sequencing of the barcode locus.
Scalable measurement of molecular responses to genetic perturbations
We have established sensitive protocols to measure small variations in fitness of cells carrying defined sequence variants via competitive growth followed by barcode sequencing. To understand how these variants lead to changes in fitness, and to more broadly assess phenotypes beyond growth we develop highly scalable screening assays based on fluorescence or sequencing.
Applying precision editing to study how genotype impacts phenotype
We apply our tools to study how traits are controlled by variation at individual nucleotides, and how these variants impact on molecular and genetic networks to shape phenotypic complexity. We are also interested in better understanding how diverse contexts alter the functional consequences of genome variants and exploiting these insights in translational applications.
The unexplored sequence diversity in natural populations is a large reservoir of untapped potential for biotechnology, synthetic biology and medicine. Using our tools we can efficiently design, engineer and screen large libraries of natural and synthetic sequence variation for advantageous properties for industrial and medical applications. Combined with our simplified protocol for microbial genome sequencing for fast and inexpensive strain validation we have the capability to generate designer biological systems with unprecedented speed, precision and efficiency.
Meet the members of the Vonesch Lab!
Sibylle VONESCH (Group Leader)
Sibylle is fascinated by the genomic and phenotypic diversity present in natural populations and loves the power of high-throughput approaches in microbes for understanding generally relevant biological principles. When not in the lab you can find her kitesurfing or running up a mountain.
Postdoc - EMBL Heidelberg, Germany
PhD - ETH Zürich, Switzerland
MSc - University of Zurich, Switzerland
Antoine DELHAYE (Postdoc)
After dabbling in biochemistry for a bit, Antoine switched focus to the intriguing field of systems biology. He loves all things microbiology and is captivated by new technologies. If you ever need to take a break from the lab and have a good night out, he's definitely a partner of choice.
PhD - UCLouvain, Belgium
MSc - Biomolecular and Cellular Engineering, UCLouvain, Belgium
Olivia OZGUC (Research Technician)
Olivia loves the creativity of synthetic biology and is passionate about developing new tools for engineering microbes. Outside of the lab she enjoys art, trying out new baking recipes, and curling up with a good book.
BSc - Biochemistry & Molecular Biology, Oregon State University, USA
Sara BACO (PhD student)
Sara has been cultivating her passion for science since she was a kid and has developed an interest in synthetic biology during her academic journey, which has led her to Belgium. She likes being in touch with nature as well as exploring cities, and after long hours in a labcoat she enjoys a cold beer while searching for a new movie to watch.
MSc - Molecular & Industrial Biotechnology, University of Bologna, Italy
Tamás TROMBITÁS (MSc student)
Tamás is inspired by the potential of genetic engineering and its expected impact on medicine, health and society. He wants to shape the future into a more sustainable and healthy life via the tools of education and science. He likes to be active and to keep himself busy with sports, music and eating delicious food.
MSc student - Cellular & Genetic Engineering, KU Leuven, Belgium
Kristof VAN ASSCHE (MSc student)
Kristof has always had a big interest in genetic engineering and its potential applications in the medical field. During his academic journey, he added to this a newfound love for statistics and is excited to be able to contribute to solving the complex problems in omics in the age of big data. During his free time he likes to be active with urban sports like breakdance or skating & occasionally likes to boulder or hike.
MSc student - Bioinformatics, KU Leuven, Belgium
Raquel MOREIRA DA SILVA (Erasmus intern)
Raquel is fascinated by genetic engineering and its potential, leading her to join the lab. From a young age all 'biothings' have captivated her, which is why you can always find her in nature or near the sea. After hours, she likes to meet friends for a good conversation and tasty dinners.
MSc student - Bioengineering, University of Porto, Portugal
Simon FIESACK (MSc student)
Simon wants to contribute to science through basic research and developing new engineering tools. Besides his interest in science, he loves tasting new things and being in nature. After a long hike, he appreciates a good beer with friends.
MSc student - Cellular & Genetic Engineering, KU Leuven, Belgium
Below is a list of our most recent publications. You can explore the full list of publications on Google Scholar.
*equal contribution #corresponding authors
Viéitez C, Busby BP, Ochoa D, Mateus A, Memon D, Galardini M, Yildiz U, Trovato M, Jawed A, Geiger AG, Oborska-Oplova M, Potel CM, Vonesch SC, Szu Tu C, Shahraz M, Stein F, Steinmetz LM, Panse VG, Noh KM, Savitski M, Typas A, Beltrao P (2021). High-throughput functional characterization of protein phosphorylation sites in yeast. Nature Biotechnology. https://doi.org/10.1038/s41587-021-01051-x
Vonesch SC#, Li S, Szu Tu C, Hennig BP, Dobrev N, Steinmetz LM# (2020). Fast and inexpensive whole genome sequencing library preparation from intact yeast cells. G3: Genes, Genomes, Genetics 11: jkaa009. https://doi.org/10.1093/g3journal/jkaa009
Vonesch SC#, Bredikhin D, Dobrev N, Villacorta L, Kleinendorst R, Cacace E, Flock J, Frank M, Jung F, Kornienko J, Mitosch K, Osuna-Lopez M, Zimmermann J, Göttig S, Hamprecht A, Kräusslich HG, Knop M, Typas A, Steinmetz LM, Benes V#, Remans K#, Krebs AR# (2020). McQ – an open-source multiplexed SARS-CoV-2 quantification platform. medRXiv. https://doi.org/10.1101/2020.12.02.20242628
Kaminski Strauss S, Schirman D, Jona G, Brooks AN, Kunjapur AM, Nguyen Ba AN, Flint A, Solt A, Mershin A, Dixit A, Yona AH, Csörgő B, Busby BP, Hennig BP, Pál C, Schraivogel D, Schultz D, Wernick DG, Agashe D, Levi D, Zabezhinsky D, Russ D, Sass E, Tamar E, Herz E, Levy ED, Church GM, Yelin I, Nachman I, Gerst JE, Georgeson JM, Adamala KP, Steinmetz LM, Rübsam M, Ralser M, Klutstein M, Desai MM, Walunjkar N, Yin N, Aharon Hefetz N, Jakimo N, Snitser O, Adini O, Kumar P, Soo Hoo Smith R, Zeidan R, Hazan R, Rak R, Kishony R, Johnson S, Nouriel S, Vonesch SC, Foster S, Dagan T, Wein T, Karydis T, Wannier TM, Stiles T, Olin-Sandoval V, Mueller WF, Bar-On YM, Dahan O, Pilpel Y (2019). Evolthon: A community endeavor to evolve lab evolution. PLoS Biology 17(3): e3000182. https://doi.org/10.1371/journal.pbio.3000182
Roy KR*, Smith JD*, Vonesch SC*, Lin G, Szu Tu C, Lederer AR, Chu A, Suresh S, Nguyen M, Horecka J, Tripathi A, Burnett WT, Morgan MA, Schulz J, Orsley KM, Wei W, Aiyar RS, Davis RW, Bankaitis VA, Haber JE, Salit ML, St.Onge RP, Steinmetz LM (2018). Multiplexed precision genome editing with trackable genomic barcodes in yeast. Nature Biotechnology 36, 512–520. https://doi.org/10.1038/nbt.4137
Baliga NS*, Björkegren JLM*, Boeke JD*, Boutros M*, Crawford NPS*, Dudley AM*, Farber CR*, Jones A*, Levey AI*, Lusis AJ*, Mak HC*, Nadeau JH*, Noyes MB*, Petretto E*, Seyfried NT*, Steinmetz LM*, Vonesch SC* (2017). The State of Systems Genetics in 2017. Cell Systems 4: 7-15.
Okada H, Ebhardt A, Vonesch SC, Aebersold R, Hafen E (2016). Proteome-wide association studies identify
biochemical modules associated with a wing size phenotype in Drosophila melanogaster. Nature
Communications 7: 12649. 10.1038/ncomms12649
Vonesch SC, Lamparter D, Mackay TFC, Bergmann S, Hafen E (2016). Genome-wide analysis reveals novel
regulators of growth in Drosophila melanogaster. PLoS Genetics 12(1): e1005616. https://doi.org/10.1371/journal.pgen.1005616