Open AccessLiquid Chromatography‐Mass Spectrometry Analysis of DNA Polymerase Reaction Products
Author(s) -
Chowdhury Goutam,
Guengerich F. Peter
Publication year - 2011
Publication title -
current protocols in nucleic acid chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.306
H-Index - 17
eISSN - 1934-9289
pISSN - 1934-9270
DOI - 10.1002/0471142700.nc0716s47
Subject(s) - primer extension , dna , chemistry , dna polymerase , nucleic acid , polymerase , mass spectrometry , chromatography , primer (cosmetics) , nucleotide , computational biology , biochemistry , biology , base sequence , gene , organic chemistry
This unit describes experimental and analytical procedures for characterizing the efficiency and fidelity of translesion DNA synthesis across various DNA damages by DNA polymerases in vitro. This procedure utilizes primer extension assays followed by LC‐MS and LC‐MS/MS analysis of the extension products. Detailed explanations for the analysis of the LC‐MS/MS data for deciphering the nucleotide sequences of the DNA fragments are also presented. This approach provides a significant improvement over conventional methods, as it allows detection of misincorporation, as well as frameshift products. Curr. Protoc. Nucleic Acid Chem . 47:7.16.1‐7.16.11. © 2011 by John Wiley & Sons, Inc.