Protocol abstract
Nature Protocols 2, - 78 - 87 (2007)
Published online: 8 February 2007 | doi:10.1038/nprot.2006.467
Subject Categories: Isolation, purification and separation | Chemical modification | Nucleic acid based molecular biology | Synthetic chemistry | Spectroscopy and structural analysis
Photochemical determination of different DNA structures
Yan Xu1, Ryu Tashiro1 & Hiroshi Sugiyama1
Abstract
The various conformations of DNA are thought to have important biological roles. Investigation of the local DNA conformational changes associated with biological events is therefore essential to an understanding of the functions of DNA. We have reported the photoreactivities of 5-halouracil in the five characteristic local DNA structures: the A, B and Z forms, protein-induced DNA kinks and the G-quadruplex form. These studies demonstrate the detailed relationships between the local DNA structures and the photochemical products of photoinduced hydrogen abstraction by the resulting uracil-5-yl radicals, and show that this photochemical method can be used to detect DNA structures. Here, we describe in detail procedures that have been developed in our laboratory for probing DNA conformations by product analysis of photoirradiated 5-halouracil–containing DNA. The protocol includes the preparation of 5-halouracil–containing DNA and the characterization of the photoproducts, and it can be completed in 2 weeks.
- Department of Chemistry, Graduate School of Science, Kyoto University, Sakyo, Kyoto 606-8502, Japan.
Correspondence to: Hiroshi Sugiyama1 e-mail: hs@kuchem.kyoto-u.ac.jp
nature-products
A-Z product listing
- 3400 DNA synthesizer(Applied Biosystems)
- 5-Bromo-2'-deoxyuridine phosphoramidite(Glen Research)
- 5-Iodo-2'-deoxyuridine phosphoramidite(Glen Research)
- Acetic acid(Sigma-Aldrich)
- Acetonitrile(Nacalai tesque)
- Ammonium formate(Wako Pure Chemical Industries)