TY - CHAP
T1 - Quantitative measurement of translesion DNA synthesis in mammalian cells
AU - Ziv, Omer
AU - Diamant, Noam
AU - Shachar, Sigal
AU - Hendel, Ayal
AU - Livneh, Zvi
PY - 2012
Y1 - 2012
N2 - 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.
AB - 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.
KW - Carcinogenesis
KW - DNA damage tolerance
KW - DNA repair
KW - Gapped plasmid
KW - Mutagenesis
KW - Site-specific DNA damage
KW - TLS
KW - Translesion DNA Synthesis
UR - http://www.scopus.com/inward/record.url?scp=84868585643&partnerID=8YFLogxK
U2 - 10.1007/978-1-61779-998-3_35
DO - 10.1007/978-1-61779-998-3_35
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C2 - 22941626
AN - SCOPUS:84868585643
SN - 9781617799976
T3 - Methods in Molecular Biology
SP - 529
EP - 542
BT - DNA Repair Protocols
PB - Humana Press Inc.
ER -