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Detection of G-Quadruplex DNA Using Primer Extension as a Tool

Kumari, Rupa and Nambiar, Mridula and Shanbagh, Shaika and Raghavan, Sathees C (2015) Detection of G-Quadruplex DNA Using Primer Extension as a Tool. In: PLOS ONE, 10 (3).

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Official URL: http://dx.doi.org/10.1371/journal.pone.0119722

Abstract

DNA sequence and structure play a key role in imparting fragility to different regions of the genome. Recent studies have shown that non-B DNA structures play a key role in causing genomic instability, apart from their physiological roles at telomeres and promoters. Structures such as G-quadruplexes, cruciforms, and triplexes have been implicated in making DNA susceptible to breakage, resulting in genomic rearrangements. Hence, techniques that aid in the easy identification of such non-B DNA motifs will prove to be very useful in determining factors responsible for genomic instability. In this study, we provide evidence for the use of primer extension as a sensitive and specific tool to detect such altered DNA structures. We have used the G-quadruplex motif, recently characterized at the BCL2 major breakpoint region as a proof of principle to demonstrate the advantages of the technique. Our results show that pause sites corresponding to the non-B DNA are specific, since they are absent when the G-quadruplex motif is mutated and their positions change in tandem with that of the primers. The efficiency of primer extension pause sites varied according to the concentration of monovalant cations tested, which support G-quadruplex formation. Overall, our results demonstrate that primer extension is a strong in vitro tool to detect non-B DNA structures such as G-quadruplex on a plasmid DNA, which can be further adapted to identify non-B DNA structures, even at the genomic level.

Item Type: Journal Article
Additional Information: Copy right for this article belongs to the PUBLIC LIBRARY SCIENCE, 1160 BATTERY STREET, STE 100, SAN FRANCISCO, CA 94111 USA
Keywords: MAJOR BREAKPOINT REGION; CHROMOSOMAL TRANSLOCATIONS; THERMOCOCCUS-LITORALIS; GENOMIC DNA; RAG COMPLEX; CANCER; STABILITY; FRAGILITY; POLYMERASE; DISEASE
Department/Centre: Division of Biological Sciences > Biochemistry
Depositing User: Id for Latest eprints
Date Deposited: 06 May 2015 05:10
Last Modified: 06 May 2015 05:10
URI: http://eprints.iisc.ac.in/id/eprint/51486

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