Nucleic Acids Research Advance Access published online on April 7, 2009
Nucleic Acids Research, doi:10.1093/nar/gkp221
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Comparative analysis of different laser systems to study cellular responses to DNA damage in mammalian cells
1Department of Biological Chemistry, 240D Med. Sci. I, School of Medicine, University of California, Irvine, CA 92697-1700, 2Beckman Laser Institute, University of California, Irvine, CA 92612 and 3Department of Bioengineering, University of California, San Diego, La Jolla, CA 92093, USA
*To whom correspondence should be addressed. Tel: +1 949 824 8215; Fax: +1 949 824 2688; Email: kyokomor{at}uci.edu
Received December 16, 2008. Revised March 19, 2009. Accepted March 20, 2009.
Proper recognition and repair of DNA damage is critical for the cell to protect its genomic integrity. Laser microirradiation ranging in wavelength from ultraviolet A (UVA) to near-infrared (NIR) can be used to induce damage in a defined region in the cell nucleus, representing an innovative technology to effectively analyze the in vivo DNA double-strand break (DSB) damage recognition process in mammalian cells. However, the damage-inducing characteristics of the different laser systems have not been fully investigated. Here we compare the nanosecond nitrogen 337 nm UVA laser with and without bromodeoxyuridine (BrdU), the nanosecond and picosecond 532 nm green second-harmonic Nd:YAG, and the femtosecond NIR 800 nm Ti:sapphire laser with regard to the type(s) of damage and corresponding cellular responses. Crosslinking damage (without significant nucleotide excision repair factor recruitment) and single-strand breaks (with corresponding repair factor recruitment) were common among all three wavelengths. Interestingly, UVA without BrdU uniquely produced base damage and aberrant DSB responses. Furthermore, the total energy required for the threshold H2AX phosphorylation induction was found to vary between the individual laser systems. The results indicate the involvement of different damage mechanisms dictated by wavelength and pulse duration. The advantages and disadvantages of each system are discussed.
Correspondence may also be addressed to Michael W. Berns. Tel: +1 949 824 7565; Fax: +1 949 824 8413; Email: mwberns{at}uci.edu
Present addresses: Jason T. Heale, Allergan, Inc. Investigative Sciences, Irvine, CA 92612, USA. Jong-Soo Kim, College of Veterinary Medicine, Seoul National University, Seoul 151-742, Korea.
The authors wish it to be known that, in their opinion, the last two authors should be regarded as joint Last Authors.
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