Andrea Callegari and Moritz Kueblbeck are Research Technicians in Jan Ellenberg Lab, EMBL. Andrea obtained his PhD in Biophysics at EPFL, Laussane. Moritz Kueblbeck worked as a Research Tecnician at the German Cancer Research Center (DKFZ) in Heidelberg.
Andrea and Moritz perform human cell line genome editing to endogenously tag proteins using CRISPR/Cas9 for different projects in the lab and with collaborators.
This webinar will describe the use of dPCR as a quick and quantitative method to measure green fluorescent protein copy number in CRISPR-edited HeLa cells. The discussion will also include an introduction to dPCR working principles and post-acquisition data analysis methods. Additionally, speakers will discuss:
- Fluorescent proteins or self-labeling tags are invaluable tools for studying protein dynamics in living cells using fluorescence microscopy. However, quantitative imaging requires physiological levels of expression of the target protein of interest (POI), especially when stoichiometric interactions of the POI need to be investigated.
- CRISPR has enabled researchers to tag virtually any target gene of interest, resulting in physiological levels of expression of the corresponding POI. However, the generation and selection of cellular clones bearing the correct edit — that is, the expected number of tagged alleles and an absence of extra integrations — requires quantitative assessment of the tag copy number.