Accedi

Helmholtz Zentrum München

3 ARTICLES PUBLISHED IN JoVE

Genetics

Real-time Analysis of Transcription Factor Binding, Transcription, Translation, and Turnover to Display Global Events During Cellular Activation

Kathrin Davari^*1, Johannes Lichti^*1, Caroline C. Friedel², Elke Glasmacher^1,3

¹Institute for Diabetes and Obesity (IDO), German Center for Diabetes Research (DZD), Helmholtz Zentrum München, ²Institute for Informatics, Ludwig-Maximilians-Universität München, ³Roche Pharma Research and Early Development, Large Molecule Research, Roche Innovation Center Penzberg

This protocol describes the combinatorial use of ChIP-seq, 4sU-seq, total RNA-seq, and ribosome profiling for cell lines and primary cells. It enables tracking changes in transcription-factor binding, de novo transcription, RNA processing, turnover and translation over time, and displaying the overall course of events in activated and/or rapidly changing cells.

Genetics

A Customizable Protocol for String Assembly gRNA Cloning (STAgR)

Christopher T. Breunig^1,2, Andrea M. Neuner^1,2, Jessica Giehrl-Schwab³, Wolfgang Wurst³, Magdalena Götz^2,4, Stefan H. Stricker^1,2

¹MCN Junior Research Group, Munich Center for Neurosciences, Ludwig Maximilian Universitat, BioMedical Center, ²Institute of Stem Cell Research, Helmholtz Zentrum, German Research Center for Environmental Health, ³Institute of Developmental Genetics, Helmholtz Zentrum, German Research Center for Environmental Health, ⁴Physiological Genomics, Ludwig Maximilian Universitat, BioMedical Center

Here, we present string assembly gRNA cloning (STAgR), a method to easily multiplex gRNA vectors for CRISPR/Cas9 approaches. STAgR makes gRNA multiplexing simple, efficient and customizable.

Neuroscience

Cryo-section Dissection of the Adult Subependymal Zone for Accurate and Deep Quantitative Proteome Analysis

Christian Friess¹, Magdalena Götz^1,2,3, Jacob Kjell^1,2,4

¹Division of Physiological Genomics, Biomedical Center, Ludwig Maximilian University of Munich, ²Institute for Stem Cell Research, Helmholtz Zentrum München, ³SYNERGY, Excellence Cluster Systems Neurology, University of Munich, ⁴Department of Clinical Neuroscience, Karolinska Institutet

Cryo-section-dissection allows fresh, frozen preparation of the largest neurogenic niche in the murine brain for deep quantitative proteome analysis. The method is precise, efficient, and causes minimal tissue perturbation. Therefore, it is ideally suited for studying the molecular microenvironment of this niche, as well as other organs, regions, and species.