Genetic code expansion for temporal controlled labeling of endogenous expressed proteins


Event details

Date 05.09.2018
Hour 16:1517:00
Speaker Prof. Matthias Meier, Helmholtz Zentrum, Muenchen
Category Conferences - Seminars

Temporal labeling of proteins is a central requisite for studying protein lifetimes, trafficking, and abundance changes in a cellular context. In this talk I will present how to combine genetic code expansion and CRISPR/Cas9 genome engineering to label specifically endogenously expressed proteins with high spatio-temporal resolution. The developed method, named SCROL (Stop-Codon-Read-thrOugh-Label), exploits the use of an orthogonal tRNA/tRNA synthetase pair in conjugation with unnatural amino acids (UAAs) to create stop codon read through events and expand protein sequences with a label. Upon timely controlled addition of a UAA, the protein sequence can be extended with a label of choice. We demonstrated the functionality of SCROL by pulse labeling of endogenous b-actin and p53 with a minimally invasive HA tag at their C-terminal ends. Synthesis rates and subcellular localization of the proteins were quantified at the single cell level with a proximity ligation assay. In contrast to existing technology, SCROL maintains the endogenous transcriptional regulatory background of the cell and does not perturb protein function in absence of the UAA.

Practical information

  • General public
  • Free


  • Prof. Fierz


  • Marie Munoz