Quantitative measurement of translesion DNA synthesis in mammalian cells

Omer Ziv, Noam Diamant, Sigal Shachar, Ayal Hendel, Zvi Livneh

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

10 Scopus citations

Abstract

Translesion DNA synthesis (TLS) is a DNA damage tolerance mechanism, in which specialized low-fidelity DNA polymerases bypass lesions that interfere with replication. This process is inherently mutagenic due to the miscoding nature of DNA lesions, but it prevents double strand breaks, genome instability, and cancer. We describe here a quantitative method for measuring TLS in mammalian cells, based on non-replicating plasmids that carry a defined and site-specific DNA lesion in a single-stranded DNA region opposite a gap. The assay is responsive to the cellular composition of TLS DNA polymerases, and TLS regulators. It can be used with a broad variety of cultured mammalian cells, and is amenable to RNAi gene silencing, making it a useful tool in the study of TLS in mammalian cells.

Original languageEnglish
Title of host publicationDNA Repair Protocols
PublisherHumana Press Inc.
Pages529-542
Number of pages14
ISBN (Print)9781617799976
DOIs
StatePublished - 2012
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume920
ISSN (Print)1064-3745

Keywords

  • Carcinogenesis
  • DNA damage tolerance
  • DNA repair
  • Gapped plasmid
  • Mutagenesis
  • Site-specific DNA damage
  • TLS
  • Translesion DNA Synthesis

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