HP-F5: Advanced Methods in Cell Biology
Type: Practical Course with Student Seminars
Date: 29. January - 16. February 2024
Locations: DKFZ Teaching Lab, TP3 Boutros Division Labs
Hosts/Supervisors: Topic A) Manuel Rodriguez-Paredes (Div. Frank Lyko)
Topic B) Kim Boonekamp and Pradhipa Manivannan, Topic C) Fillip Port (all Div. Michael Boutros)
Organizer contact: Ulrike Hardeland (Div. Boutros) u.hardeland@dkfz.de
Topics:
- A) Role of UV radiation in human skin epigenetic deregulation leading to cancer
- B) Imaged-based screening to study gene function and drug sensitivity
- C) Tissue-specific CRISPR/Cas genome engineering in vivo
Content:
general: Cell culture, genomic DNA/RNA isolation, RNA retrotranscription (RT), immunofluorescence microscopy, Western blotting, quantitative RT-PCR, cDNA transfection, DNA bisulfite conversion and methylation-sensitive PCR (MSP), RNAi screening, high-throughput data analysis
details for topic A): DNA methylation at gene promoters and other genomic regulatory elements is probably the most studied epigenetic modification. Using cell lines as well as students' own skin tissue, in this part of the course we will investigate how UV light causes DNA damage and affects the methylome of human epidermis, leading to gene expression changes that ultimately favor skin cancer development.
details for topic B): Imaged-based screening has become an important method for the characterization of gene function and drug effect. As part of the course, participants will learn how to perform image-based screenings in mammalian cell lines and how to analyze these screens. We will perform a small image-based screening assay on 2D cell lines to address chemical-genetic interactions (CGI) in mammalian cells relevant to cancer therapies. The course will include hands-on practice in data analysis of high-content profiling experiments using state of the art computational methods.
details for topic C): CRISPR/Cas genome engineering permits targeted genome manipulations with unprecedented ease and precision. We have developed state-of the art CRISPR systems based on Cas9, Cas12a as well as prime- and base-editing tools that permit conditional genome engineering within a multicellular organism. Using Drosophila melanogaster as a model, participants will manipulate candidate genes in a subset of epithelial cells and analyze the resulting phenotypes by high-resolution microscopy and sequencing based readouts.