Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments

Joel Tellinghuisen; Andrej-Nikolai Spiess

doi:10.1021/acs.analchem.5b00077

Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments

Standard

Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments. / Tellinghuisen, Joel; Spiess, Andrej-Nikolai.

in: ANAL CHEM, Jahrgang 87, Nr. 3, 03.02.2015, S. 1889-95.

Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung

Harvard

Tellinghuisen, J & Spiess, A-N 2015, 'Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments', ANAL CHEM, Jg. 87, Nr. 3, S. 1889-95. https://doi.org/10.1021/acs.analchem.5b00077

APA

Tellinghuisen, J., & Spiess, A-N. (2015). Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments. ANAL CHEM, 87(3), 1889-95. https://doi.org/10.1021/acs.analchem.5b00077

Vancouver

Tellinghuisen J, Spiess A-N. Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments. ANAL CHEM. 2015 Feb 3;87(3):1889-95. https://doi.org/10.1021/acs.analchem.5b00077

Bibtex

@article{fa358c66226e47189b348c5505076b0a,

title = "Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments",

abstract = "The quantification cycle (Cq) is widely used for calibration in real-time quantitative polymerase chain reaction (qPCR), to estimate the initial amount, or copy number (N0), of the target DNA. Cq may be defined several ways, including the cycle where the detected fluorescence achieves a prescribed threshold level. For all methods of defining Cq, the standard deviation from replicate experiments is typically much greater than the estimated standard errors from the least-squares fits used to obtain Cq. For moderate-to-large copy number (N0 > 10(2)), pipet volume uncertainty and variability in the amplification efficiency (E) likely account for most of the excess variance in Cq. For small N0, the dispersion of Cq is determined by the Poisson statistics of N0, which means that N0 can be estimated directly from the variance of Cq. The estimation precision is determined by the statistical properties of χ(2), giving a relative standard deviation of ∼(2/n)(1/2), where n is the number of replicates, for example, a 20% standard deviation in N0 from 50 replicates.",

keywords = "Analysis of Variance, Gene Dosage, Least-Squares Analysis, Real-Time Polymerase Chain Reaction",

author = "Joel Tellinghuisen and Andrej-Nikolai Spiess",

year = "2015",

month = feb,

day = "3",

doi = "10.1021/acs.analchem.5b00077",

language = "English",

volume = "87",

pages = "1889--95",

journal = "ANAL CHEM",

issn = "0003-2700",

publisher = "American Chemical Society",

number = "3",

}

RIS

TY - JOUR

T1 - Absolute copy number from the statistics of the quantification cycle in replicate quantitative polymerase chain reaction experiments

AU - Tellinghuisen, Joel

AU - Spiess, Andrej-Nikolai

PY - 2015/2/3

Y1 - 2015/2/3

N2 - The quantification cycle (Cq) is widely used for calibration in real-time quantitative polymerase chain reaction (qPCR), to estimate the initial amount, or copy number (N0), of the target DNA. Cq may be defined several ways, including the cycle where the detected fluorescence achieves a prescribed threshold level. For all methods of defining Cq, the standard deviation from replicate experiments is typically much greater than the estimated standard errors from the least-squares fits used to obtain Cq. For moderate-to-large copy number (N0 > 10(2)), pipet volume uncertainty and variability in the amplification efficiency (E) likely account for most of the excess variance in Cq. For small N0, the dispersion of Cq is determined by the Poisson statistics of N0, which means that N0 can be estimated directly from the variance of Cq. The estimation precision is determined by the statistical properties of χ(2), giving a relative standard deviation of ∼(2/n)(1/2), where n is the number of replicates, for example, a 20% standard deviation in N0 from 50 replicates.

AB - The quantification cycle (Cq) is widely used for calibration in real-time quantitative polymerase chain reaction (qPCR), to estimate the initial amount, or copy number (N0), of the target DNA. Cq may be defined several ways, including the cycle where the detected fluorescence achieves a prescribed threshold level. For all methods of defining Cq, the standard deviation from replicate experiments is typically much greater than the estimated standard errors from the least-squares fits used to obtain Cq. For moderate-to-large copy number (N0 > 10(2)), pipet volume uncertainty and variability in the amplification efficiency (E) likely account for most of the excess variance in Cq. For small N0, the dispersion of Cq is determined by the Poisson statistics of N0, which means that N0 can be estimated directly from the variance of Cq. The estimation precision is determined by the statistical properties of χ(2), giving a relative standard deviation of ∼(2/n)(1/2), where n is the number of replicates, for example, a 20% standard deviation in N0 from 50 replicates.

KW - Analysis of Variance

KW - Gene Dosage

KW - Least-Squares Analysis

KW - Real-Time Polymerase Chain Reaction

U2 - 10.1021/acs.analchem.5b00077

DO - 10.1021/acs.analchem.5b00077

M3 - SCORING: Journal article

C2 - 25582662

VL - 87

SP - 1889

EP - 1895

JO - ANAL CHEM

JF - ANAL CHEM

SN - 0003-2700

IS - 3

ER -