Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number

Abstract Recently, in food safety and various other fields, qualitative and quantitative gene analysis using real-time polymerase chain reaction (PCR) method has become increasingly popular. The limit of detection (LOD) and quantifiable range for these measurements depends on the range and precision...

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Autores principales:	Unoh Ki, Takeru Suzuki, Satoshi Nakazawa, Yuuki Yonekawa, Kazuki Watanabe, Michie Hashimoto, Shigeo Hatada, Hirotaka Unno
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/37017f5d07c74d3abb547ec4bdde3f85
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spelling	oai:doaj.org-article:37017f5d07c74d3abb547ec4bdde3f852021-12-02T15:03:05ZEvaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number10.1038/s41598-021-90959-02045-2322https://doaj.org/article/37017f5d07c74d3abb547ec4bdde3f852021-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-90959-0https://doaj.org/toc/2045-2322Abstract Recently, in food safety and various other fields, qualitative and quantitative gene analysis using real-time polymerase chain reaction (PCR) method has become increasingly popular. The limit of detection (LOD) and quantifiable range for these measurements depends on the range and precision of DNA calibrators’ concentrations. Low-copy-number nucleic acid reference materials with low uncertainty produced by an inkjet system have been developed to allow for precise measurements in a low-copy-number region. However, when using a calibrator with a low copy number near one, the copy number distribution is asymmetric. Consequently, the confidence intervals of estimated copy numbers can include negative values when conventional methods of uncertainty estimation are used. A negative confidence interval is irrelevant in the context of copy number, which is always positive value or zero. Here, we propose a method to evaluate the uncertainty of real-time PCR measurements with representative values and an asymmetric 95% confidence interval. Moreover, we use the proposed method for the actual calculation of uncertainty of real-time PCR measurement results for low-copy-number DNA samples and demonstrate that the proposed method can evaluate the precision of real-time PCR measurements more appropriately in a low-copy-number region.Unoh KiTakeru SuzukiSatoshi NakazawaYuuki YonekawaKazuki WatanabeMichie HashimotoShigeo HatadaHirotaka UnnoNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-16 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Unoh Ki Takeru Suzuki Satoshi Nakazawa Yuuki Yonekawa Kazuki Watanabe Michie Hashimoto Shigeo Hatada Hirotaka Unno Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
description	Abstract Recently, in food safety and various other fields, qualitative and quantitative gene analysis using real-time polymerase chain reaction (PCR) method has become increasingly popular. The limit of detection (LOD) and quantifiable range for these measurements depends on the range and precision of DNA calibrators’ concentrations. Low-copy-number nucleic acid reference materials with low uncertainty produced by an inkjet system have been developed to allow for precise measurements in a low-copy-number region. However, when using a calibrator with a low copy number near one, the copy number distribution is asymmetric. Consequently, the confidence intervals of estimated copy numbers can include negative values when conventional methods of uncertainty estimation are used. A negative confidence interval is irrelevant in the context of copy number, which is always positive value or zero. Here, we propose a method to evaluate the uncertainty of real-time PCR measurements with representative values and an asymmetric 95% confidence interval. Moreover, we use the proposed method for the actual calculation of uncertainty of real-time PCR measurement results for low-copy-number DNA samples and demonstrate that the proposed method can evaluate the precision of real-time PCR measurements more appropriately in a low-copy-number region.
format	article
author	Unoh Ki Takeru Suzuki Satoshi Nakazawa Yuuki Yonekawa Kazuki Watanabe Michie Hashimoto Shigeo Hatada Hirotaka Unno
author_facet	Unoh Ki Takeru Suzuki Satoshi Nakazawa Yuuki Yonekawa Kazuki Watanabe Michie Hashimoto Shigeo Hatada Hirotaka Unno
author_sort	Unoh Ki
title	Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
title_short	Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
title_full	Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
title_fullStr	Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
title_full_unstemmed	Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
title_sort	evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/37017f5d07c74d3abb547ec4bdde3f85
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_version_	1718389070161248256

Evaluation method for asymmetric uncertainty of quantitative polymerase chain reaction measurements of deoxyribonucleic acids with low copy number

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