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Use of 5-Ethynyl-2′-Deoxyuridine Labelling and Flow Cytometry to Study Cell Cycle-Dependent Regulation of Human Cytomegalovirus Gene Expression

  • Lüder WiebuschEmail author
  • Christian Hagemeier
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1119)

Abstract

The cell cycle position at the time of infection has a profound influence on human cytomegalovirus (HCMV) gene expression and therefore needs consideration in the design and control of HCMV experiments. While G0/G1 cells support the immediate onset of viral transcription, cells progressing through the S and G2 cell cycle phases prevent HCMV from entering the lytic replication cycle. Here, we provide two fast and reliable protocols that allow one to determine the cell cycle distribution of the designated host cells and monitor viral protein expression as a function of the cell cycle state. Both protocols make use of the thymidine analogue 5-ethynyl-2′-deoxyuridine and “click” chemistry to label HCMV-non-permissive S phase cells in a gentle and sensitive way.

Key words

Cell cycle Cytomegalovirus EdU labelling Flow cytometry HCMV antibodies Immediate early genes Propidium iodide staining 

Notes

Acknowledgement

This work has been supported by the DFG grant WI2043/3-1 to L.W. and C.H.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Labor für Pädiatrische MolekularbiologieCharité UniversitätsmedizinBerlinGermany
  2. 2.Labor für Pädiatrische MolekularbiologieCharité UniversitätsmedizinBerlinGermany

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