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Simplifying the Detection of Surface Presentation Levels in Yeast Surface Display by Intracellular tGFP Expression

  • Steffen C. Hinz
  • Adrian Elter
  • Julius Grzeschik
  • Jan Habermann
  • Bastian Becker
  • Harald KolmarEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2070)

Abstract

Yeast surface display (YSD) is an ultra-high throughput method used in protein engineering. Protein-protein interactions as well as surface presentation on the yeast cell surface are verified through fluorophore-conjugated labeling agents.

In this chapter we describe an improved setup for full-length surface presentation detection. To this end, we used a single open reading frame (ORF) encoding for the protein to be displayed and a 2A sequence and tGFP for an intracellular fluorescence signal. The 2A sequence allows the simultaneous generation of two separate proteins from the same ORF through ribosomal skipping. The entangled expression of the POI on the yeast surface and intracellular tGFP obviates the need for fluorescent staining steps.

Key words

T2A 2A peptide Yeast surface display GFP Ribosomal skipping 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Steffen C. Hinz
    • 1
  • Adrian Elter
    • 1
  • Julius Grzeschik
    • 1
  • Jan Habermann
    • 1
  • Bastian Becker
    • 1
  • Harald Kolmar
    • 1
    Email author
  1. 1.Institute for Organic Chemistry and BiochemistryTechnische Universität DarmstadtDarmstadtGermany

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