NanoHDA: A nanoparticle-assisted isothermal amplification technique for genotyping assays

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• 作者：Abootaleb Sedighi ; Christopher Oberc ; Vicki Whitehall ; Paul C. H. Li
• 关键词：gold nanoparticle ; isothermal DNA amplification ; helicase ; genotyping assay ; colorectal cancer ; single nucleotide polymorphism ; KRAS
• 刊名：Nano Research
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：10
• 期：1
• 页码：12-21
• 全文大小：
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Nanotechnology; Materials Science, general; Atomic/Molecular Structure and Spectra; Condensed Matter Physics; Biotechnology; Biomedicine, general;
• 出版者：Tsinghua University Press
• ISSN：1998-0000
• 卷排序：10

文摘

Isothermal methods, such as helicase-dependent amplification (HDA), have an advantage over polymerase chain reaction for DNA amplification owing to their ease of operation. Here, we developed a new HDA method that is nanoparticle-assisted, termed nanoHDA. This method uses gold nanoparticles (AuNPs) to improve the sensitivity and specificity of the isothermal method. In HDA, the denaturation of DNA templates is mediated by helicases, but this method is limited by the low denaturation efficiency of helicases. In this report, AuNPs with preferential affinity for single-stranded DNA (ssDNA) were utilized to improve the denaturation efficiency of helicases. The same affinity property of nanoparticles can also enhance specificity by suppressing primer-dimer formation. This nanoHDA method was employed to genotype the KRAS gene in genomic DNA samples from colorectal cancer patients, as achieved by the hybridization of nanoHDA amplicons using the NanoBioArray chip.