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A colorimetric method for point mutation detection using high-fidelity DNA ligase

The present study reported proof-of-principle for a genotyping assay approach that can detect single nucleotide polymorphisms (SNPs) through the gold nanoparticle assembly and the ligase reaction. By incorporating the high-fidelity DNA ligase (Tth DNA ligase) into the allele-specific ligation-based...

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Autores principales: Li, Jishan, Chu, Xia, Liu, Yali, Jiang, Jian-Hui, He, Zhimin, Zhang, Zhiwei, Shen, Guoli, Yu, Ru-Qin
Formato: Texto
Lenguaje:English
Publicado: Oxford University Press 2005
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1275593/
https://www.ncbi.nlm.nih.gov/pubmed/16257979
http://dx.doi.org/10.1093/nar/gni163
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author Li, Jishan
Chu, Xia
Liu, Yali
Jiang, Jian-Hui
He, Zhimin
Zhang, Zhiwei
Shen, Guoli
Yu, Ru-Qin
author_facet Li, Jishan
Chu, Xia
Liu, Yali
Jiang, Jian-Hui
He, Zhimin
Zhang, Zhiwei
Shen, Guoli
Yu, Ru-Qin
author_sort Li, Jishan
collection PubMed
description The present study reported proof-of-principle for a genotyping assay approach that can detect single nucleotide polymorphisms (SNPs) through the gold nanoparticle assembly and the ligase reaction. By incorporating the high-fidelity DNA ligase (Tth DNA ligase) into the allele-specific ligation-based gold nanoparticle assembly, this assay provided a convenient yet powerful colorimetric detection that enabled a straightforward single-base discrimination without the need of precise temperature control. Additionally, the ligase reaction can be performed at a relatively high temperature, which offers the benefit for mitigating the non-specific assembly of gold nanoparticles induced by interfering DNA strands. The assay could be implemented via three steps: a hybridization reaction that allowed two gold nanoparticle-tagged probes to hybrid with the target DNA strand, a ligase reaction that generates the ligation between perfectly matched probes while no ligation occurred between mismatched ones and a thermal treatment at a relatively high temperature that discriminate the ligation of probes. When the reaction mixture was heated to denature the formed duplex, the purple color of the perfect-match solution would not revert to red, while the mismatch gave a red color as the assembled gold nanoparticles disparted. The present approach has been demonstrated with the identification of a single-base mutation in codon 12 of a K-ras oncogene that is of significant value for colorectal cancers diagnosis, and the wild-type and mutant type were successfully scored. To our knowledge, this was the first report concerning SNP detection based on the ligase reaction and the gold nanoparticle assembly. Owing to its ease of operation and high specificity, it was expected that the proposed procedure might hold great promise in practical clinical diagnosis of gene-mutant diseases.
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spelling pubmed-12755932005-11-01 A colorimetric method for point mutation detection using high-fidelity DNA ligase Li, Jishan Chu, Xia Liu, Yali Jiang, Jian-Hui He, Zhimin Zhang, Zhiwei Shen, Guoli Yu, Ru-Qin Nucleic Acids Res Methods Online The present study reported proof-of-principle for a genotyping assay approach that can detect single nucleotide polymorphisms (SNPs) through the gold nanoparticle assembly and the ligase reaction. By incorporating the high-fidelity DNA ligase (Tth DNA ligase) into the allele-specific ligation-based gold nanoparticle assembly, this assay provided a convenient yet powerful colorimetric detection that enabled a straightforward single-base discrimination without the need of precise temperature control. Additionally, the ligase reaction can be performed at a relatively high temperature, which offers the benefit for mitigating the non-specific assembly of gold nanoparticles induced by interfering DNA strands. The assay could be implemented via three steps: a hybridization reaction that allowed two gold nanoparticle-tagged probes to hybrid with the target DNA strand, a ligase reaction that generates the ligation between perfectly matched probes while no ligation occurred between mismatched ones and a thermal treatment at a relatively high temperature that discriminate the ligation of probes. When the reaction mixture was heated to denature the formed duplex, the purple color of the perfect-match solution would not revert to red, while the mismatch gave a red color as the assembled gold nanoparticles disparted. The present approach has been demonstrated with the identification of a single-base mutation in codon 12 of a K-ras oncogene that is of significant value for colorectal cancers diagnosis, and the wild-type and mutant type were successfully scored. To our knowledge, this was the first report concerning SNP detection based on the ligase reaction and the gold nanoparticle assembly. Owing to its ease of operation and high specificity, it was expected that the proposed procedure might hold great promise in practical clinical diagnosis of gene-mutant diseases. Oxford University Press 2005 2005-10-27 /pmc/articles/PMC1275593/ /pubmed/16257979 http://dx.doi.org/10.1093/nar/gni163 Text en © The Author 2005. Published by Oxford University Press. All rights reserved
spellingShingle Methods Online
Li, Jishan
Chu, Xia
Liu, Yali
Jiang, Jian-Hui
He, Zhimin
Zhang, Zhiwei
Shen, Guoli
Yu, Ru-Qin
A colorimetric method for point mutation detection using high-fidelity DNA ligase
title A colorimetric method for point mutation detection using high-fidelity DNA ligase
title_full A colorimetric method for point mutation detection using high-fidelity DNA ligase
title_fullStr A colorimetric method for point mutation detection using high-fidelity DNA ligase
title_full_unstemmed A colorimetric method for point mutation detection using high-fidelity DNA ligase
title_short A colorimetric method for point mutation detection using high-fidelity DNA ligase
title_sort colorimetric method for point mutation detection using high-fidelity dna ligase
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1275593/
https://www.ncbi.nlm.nih.gov/pubmed/16257979
http://dx.doi.org/10.1093/nar/gni163
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