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RNA: a method to specifically inhibit PCR amplification of known members of a multigene family by degenerate primers

Nucleic Acids Research
Oxford University Press
Publication Date
  • Nar Methods Online
  • Biology


OP-NARE131233 1..16 Novel method for site-specific induction of oxidative DNA damage reveals differences in recruitment of repair proteins to heterochromatin and euchromatin Li Lan1,2,*, Satoshi Nakajima1,2, Leizhen Wei1,2, Luxi Sun2,3, Ching-Lung Hsieh1,2, Robert W. Sobol2,4,5, Marcel Bruchez6, Bennett Van Houten2,4, Akira Yasui7 and Arthur S. Levine1,2 1Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA, 2University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, 5117 Centre Avenue, Pittsburgh, PA 15213, USA, 3School of Medicine, Tsinghua University, No.1 Tsinghua Yuan, Haidian District, Beijing 100084, People’s Republic of China, 4Department of Pharmacology & Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA, 5Department of Human Genetics, University of Pittsburgh Graduate School of Public Health, Pittsburgh, PA 15213, USA, 6Department of Chemistry and Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213, USA and 7Division of Dynamic Proteome, Institute of Development, Aging, and Cancer, Tohoku University, Seiryomachi 4-1, Sendai 980-8575, Japan Received August 10, 2013; Revised November 5, 2013; Accepted November 6, 2013 ABSTRACT Reactive oxygen species (ROS)-induced DNA damage is repaired by the base excision repair pathway. However, the effect of chromatin structure on BER protein recruitment to DNA damage sites in living cells is poorly understood. To address this problem, we developed a method to specifically produce ROS-induced DNA damage by fusing KillerRed (KR), a light-stimulated ROS-inducer, to a tet-repressor (tetR-KR) or a transcription activator (TA-KR). TetR-KR or TA-KR, bound to a TRE cassette (�90kb) integrated at a defined genomic locus in U2OS cells, was used to induce ROS damage in hetero- or euchromatin, respectively. We found that DNA glycosylases were efficiently recruited to DNA damage in h

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