A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas
We have characterized a novel nuclease from the Kamchatka crab, designated duplex-specific nuclease (DSN). DSN displays a strong preference for cleaving double-stranded DNA and DNA in DNA-RNA hybrid duplexes, compared to single-stranded DNA. Moreover, the cleavage rate of short, perfectly matched DN...
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ftpubmed:oai:pubmedcentral.nih.gov:187582 2023-05-15T16:59:15+02:00 A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas Shagin, Dmitry A. Rebrikov, Denis V. Kozhemyako, Valery B. Altshuler, Ilia M. Shcheglov, Alex S. Zhulidov, Pavel A. Bogdanova, Ekaterina A. Staroverov, Dmitry B. Rasskazov, Valery A. Lukyanov, Sergey 2002-12 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC187582 http://www.ncbi.nlm.nih.gov/pubmed/12466298 https://doi.org/10.1101/gr.547002 en eng Cold Spring Harbor Laboratory Press http://www.ncbi.nlm.nih.gov/pmc/articles/PMC187582 http://www.ncbi.nlm.nih.gov/pubmed/12466298 http://dx.doi.org/10.1101/gr.547002 Copyright © 2002, Cold Spring Harbor Laboratory Press Methods Text 2002 ftpubmed https://doi.org/10.1101/gr.547002 2013-08-29T13:34:51Z We have characterized a novel nuclease from the Kamchatka crab, designated duplex-specific nuclease (DSN). DSN displays a strong preference for cleaving double-stranded DNA and DNA in DNA-RNA hybrid duplexes, compared to single-stranded DNA. Moreover, the cleavage rate of short, perfectly matched DNA duplexes by this enzyme is essentially higher than that for nonperfectly matched duplexes of the same length. Thus, DSN differentiates between one-nucleotide variations in DNA. We developed a novel assay for single nucleotide polymorphism (SNP) detection based on this unique property, termed “duplex-specific nuclease preference” (DSNP). In this innovative assay, the DNA region containing the SNP site is amplified and the PCR product mixed with signal probes (FRET-labeled short sequence-specific oligonucleotides) and DSN. During incubation, only perfectly matched duplexes between the DNA template and signal probe are cleaved by DSN to generate sequence-specific fluorescence. The use of FRET-labeled signal probes coupled with the specificity of DSN presents a simple and efficient method for detecting SNPs. We have employed the DSNP assay for the typing of SNPs in methyltetrahydrofolate reductase, prothrombin and p53 genes on homozygous and heterozygous genomic DNA. Text Kamchatka Kamchatka crab PubMed Central (PMC) Genome Research 12 12 1935 1942 |
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Methods Shagin, Dmitry A. Rebrikov, Denis V. Kozhemyako, Valery B. Altshuler, Ilia M. Shcheglov, Alex S. Zhulidov, Pavel A. Bogdanova, Ekaterina A. Staroverov, Dmitry B. Rasskazov, Valery A. Lukyanov, Sergey A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
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Methods |
description |
We have characterized a novel nuclease from the Kamchatka crab, designated duplex-specific nuclease (DSN). DSN displays a strong preference for cleaving double-stranded DNA and DNA in DNA-RNA hybrid duplexes, compared to single-stranded DNA. Moreover, the cleavage rate of short, perfectly matched DNA duplexes by this enzyme is essentially higher than that for nonperfectly matched duplexes of the same length. Thus, DSN differentiates between one-nucleotide variations in DNA. We developed a novel assay for single nucleotide polymorphism (SNP) detection based on this unique property, termed “duplex-specific nuclease preference” (DSNP). In this innovative assay, the DNA region containing the SNP site is amplified and the PCR product mixed with signal probes (FRET-labeled short sequence-specific oligonucleotides) and DSN. During incubation, only perfectly matched duplexes between the DNA template and signal probe are cleaved by DSN to generate sequence-specific fluorescence. The use of FRET-labeled signal probes coupled with the specificity of DSN presents a simple and efficient method for detecting SNPs. We have employed the DSNP assay for the typing of SNPs in methyltetrahydrofolate reductase, prothrombin and p53 genes on homozygous and heterozygous genomic DNA. |
format |
Text |
author |
Shagin, Dmitry A. Rebrikov, Denis V. Kozhemyako, Valery B. Altshuler, Ilia M. Shcheglov, Alex S. Zhulidov, Pavel A. Bogdanova, Ekaterina A. Staroverov, Dmitry B. Rasskazov, Valery A. Lukyanov, Sergey |
author_facet |
Shagin, Dmitry A. Rebrikov, Denis V. Kozhemyako, Valery B. Altshuler, Ilia M. Shcheglov, Alex S. Zhulidov, Pavel A. Bogdanova, Ekaterina A. Staroverov, Dmitry B. Rasskazov, Valery A. Lukyanov, Sergey |
author_sort |
Shagin, Dmitry A. |
title |
A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
title_short |
A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
title_full |
A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
title_fullStr |
A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
title_full_unstemmed |
A Novel Method for SNP Detection Using a New Duplex-Specific Nuclease From Crab Hepatopancreas |
title_sort |
novel method for snp detection using a new duplex-specific nuclease from crab hepatopancreas |
publisher |
Cold Spring Harbor Laboratory Press |
publishDate |
2002 |
url |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC187582 http://www.ncbi.nlm.nih.gov/pubmed/12466298 https://doi.org/10.1101/gr.547002 |
genre |
Kamchatka Kamchatka crab |
genre_facet |
Kamchatka Kamchatka crab |
op_relation |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC187582 http://www.ncbi.nlm.nih.gov/pubmed/12466298 http://dx.doi.org/10.1101/gr.547002 |
op_rights |
Copyright © 2002, Cold Spring Harbor Laboratory Press |
op_doi |
https://doi.org/10.1101/gr.547002 |
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Genome Research |
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12 |
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12 |
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1935 |
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1942 |
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1766051487189827584 |