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Locus-Specific Knock-In of a Degradable Tag for Target Validation Studies

  • Matthias Brand
  • Georg E. WinterEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1953)

Abstract

Small molecule-induced targeted protein degradation is a powerful approach for drug target validation given its selectivity, high kinetic resolution, dose dependency, and reversibility. Out of the several methods that have been reported so far, the 12 kDa degradation tag (dTAG) system has the advantage of hijacking a degradation machinery that is ubiquitously expressed in all human tissues. Therefore it is independent of expressing additional, exogenous factors and additionally permits target validation in vivo. Here, we describe the protocol for generation and validation of clones harboring knock-in of a selectable dTAG cassette at the endogenous locus of proteins of interest using the near-haploid cell line KBM7.

Key words

dTAG PROTACs Targeted protein degradation Genome engineering Target validation 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.CeMM Research Center for Molecular Medicine of the Austrian Academy of SciencesViennaAustria

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