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SUMMARY:Genetic code expansion for temporal controlled labeling of endogen
 ous expressed proteins
DTSTART:20180905T161500
DTEND:20180905T170000
DTSTAMP:20260405T200440Z
UID:60e8e23aa794667971c54276299db771a713d57340b8874ca716f8bb
CATEGORIES:Conferences - Seminars
DESCRIPTION:Prof. Matthias Meier\, Helmholtz Zentrum\, Muenchen\nTemporal 
 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 geno
 me 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 sy
 nthetase pair in conjugation with unnatural amino acids (UAAs) to create s
 top codon read through events and expand protein sequences with a label. U
 pon timely controlled addition of a UAA\, the protein sequence can be exte
 nded 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 localizatio
 n of the proteins were quantified at the single cell level with a proximit
 y 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.\n 
LOCATION:BCH 4310 https://plan.epfl.ch/?room==BCH%204310
STATUS:CONFIRMED
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