Sensitive detection of DNA polymorphisms by the serial invasive signal amplification reaction.
Jeff G. Hall,Peggy S. Eis,Scott M. Law,Luis P. Reynaldo,James R. Prudent,David J. Marshall,Hatim T. Allawi,Andrea L. Mast,James E. Dahlberg,Robert W. Kwiatkowski,Monika de Arruda,Bruce Neri,Victor I. Lyamichev +12 more
TLDR
The invasive signal amplification reaction is described as a method that couples two invasive reactions into a serial isothermal homogeneous assay using fluorescence resonance energy transfer detection and a simple kinetic model of the assay is described.Abstract:
The invasive signal amplification reaction has been previously developed for quantitative detection of nucleic acids and discrimination of single-nucleotide polymorphisms. Here we describe a method that couples two invasive reactions into a serial isothermal homogeneous assay using fluorescence resonance energy transfer detection. The serial version of the assay generates more than 10(7) reporter molecules for each molecule of target DNA in a 4-h reaction; this sensitivity, coupled with the exquisite specificity of the reaction, is sufficient for direct detection of less than 1,000 target molecules with no prior target amplification. Here we present a kinetic analysis of the parameters affecting signal and background generation in the serial invasive signal amplification reaction and describe a simple kinetic model of the assay. We demonstrate the ability of the assay to detect as few as 600 copies of the methylene tetrahydrofolate reductase gene in samples of human genomic DNA. We also demonstrate the ability of the assay to discriminate single base differences in this gene by using 20 ng of human genomic DNA.read more
Citations
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Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems.
TL;DR: A snapshot of the state-of-the-art in real-time RT-PCR is provided and some of the problems associated with interpreting results that are numerical and lend themselves to statistical analysis, yet whose accuracy is significantly affected by reagent and operator variability are described.
Journal ArticleDOI
Isothermal Amplification of Nucleic Acids
TL;DR: This review provides a comprehensive overview of the isothermal amplification of nucleic acids encompassing work published in the past two decades including applications in bioanalysis, diagnostics, nanotechnology, materials science, and device integration.
Journal ArticleDOI
Accessing genetic variation: genotyping single nucleotide polymorphisms
TL;DR: The hope that single nucleotide polymorphisms will allow genes that underlie complex disease to be identified, together with progress in identifying large sets ofSNPs, are the driving forces behind intense efforts to establish the technology for large-scale analysis of SNPs.
Journal ArticleDOI
Methods for genotyping single nucleotide polymorphisms.
TL;DR: In this review, the current status of SNP genotyping is discussed in terms of the mechanisms of allelic discrimination, the reaction formats, and the detection modalities.
Journal ArticleDOI
SNP Genotyping: Technologies and Biomedical Applications
Sobin Kim,Ashish Misra +1 more
TL;DR: This article presents a review of SNP genotyping techniques and examines their principles of genotype determination in terms of allele differentiation strategies and detection methods.
References
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Book ChapterDOI
Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction.
Kary B. Mullis,Fred A. Faloona +1 more
TL;DR: A method whereby a nucleic acid sequence can be exponentially amplified in vitro is described in the chapter, and the possibility of utilizing a heat-stable DNA polymerase is explored so as to avoid the need for addition of new enzyme after each cycle of thermal denaturation.
Journal ArticleDOI
A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase
P. Frosst,Henk J. Blom,Renate Milos,Philippe Goyette,Christal A. Sheppard,Rowena G. Matthews,G. J.H. Boers,M. den Heijer,Leo A. J. Kluijtmans,L.P.W.J. van den Heuvel,Rima Rozen +10 more
TL;DR: This work has identified a common mutation in MTHFR which alters a highly-conserved amino acid; the substitution occurs at a frequency of approximately 38% of unselected chromosomes and may represent an important genetic risk factor in vascular disease.
Journal ArticleDOI
Rate of depurination of native deoxyribonucleic acid.
Tomas Lindahl,Barbro Nyberg +1 more
Journal ArticleDOI
Typing Hepatitis B Virus by Homology in Nucleotide Sequence: Comparison of Surface Antigen Subtypes
Hiroaki Okamoto,Fumio Tsuda,Hiroshi Sakugawa,Retno I. Sastrosoewignjo,Mitsunobu Imai,Yuzo Miyakawa,Makoto Mayumi +6 more
TL;DR: The results indicate that the four major antigenically defined subtypes of envelope polypeptide do not reflect true genotypic variation of HBV and can be induced by an A----G point mutation at nucleotides 365 and 479 in the S gene.
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Thermodynamics and NMR of Internal G·T Mismatches in DNA
Hatim T. Allawi,John SantaLucia +1 more
TL;DR: Analysis of imino proton chemical shifts show that structural perturbations from the G.T mismatches are highly localized and form stable hydrogen-bonded wobble pairs in diverse contexts.
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