Dataset Information

Real-time relative qPCR without reference to control samples and estimation of run-specific PCR parameters from run-internal mini-standard curves.

ABSTRACT:

Background

Real-Time quantitative PCR is an important tool in research and clinical settings. Here, we describe two new approaches that broaden the scope of real-time quantitative PCR; namely, run-internal mini standard curves (RIMS) and direct real-time relative quantitative PCR (drqPCR). RIMS are an efficient alternative to traditional standard curves and provide both run-specific and target-specific estimates of PCR parameters. The drqPCR enables direct estimation of target ratios without reference to conventional control samples.

Methodology/principal findings

In this study, we compared RIMS-based drqPCR with classical quantifications based on external standard curves and the "comparative Ct method". Specifically, we used a raw real-time PCR dataset as the basis for more than two-and-a-half million simulated quantifications with various user-defined conditions. Compared with classical approaches, we found that RIMS-based drqPCR provided superior precision and comparable accuracy.

Conclusions/significance

The obviation of referencing to control samples is attractive whenever unpaired samples are quantified. This may be in clinical and research settings; for instance, studies on chimerism, TREC quantifications, copy number variations etc. Also, lab-to-lab comparability can be greatly simplified.

SUBMITTER: Bernth Jensen JM

PROVIDER: S-EPMC2908630 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Publications

Real-time relative qPCR without reference to control samples and estimation of run-specific PCR parameters from run-internal mini-standard curves.

Bernth Jensen Jens Magnus JM Petersen Mikkel Steen MS Stegger Marc M Østergaard Lars J LJ Møller Bjarne K BK

PloS one 20100722 7

<h4>Background</h4>Real-Time quantitative PCR is an important tool in research and clinical settings. Here, we describe two new approaches that broaden the scope of real-time quantitative PCR; namely, run-internal mini standard curves (RIMS) and direct real-time relative quantitative PCR (drqPCR). RIMS are an efficient alternative to traditional standard curves and provide both run-specific and target-specific estimates of PCR parameters. The drqPCR enables direct estimation of target ratios wit ...[more]

PMID: 20661435

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Dataset Information

Real-time relative qPCR without reference to control samples and estimation of run-specific PCR parameters from run-internal mini-standard curves.

Background

Methodology/principal findings

Conclusions/significance

Publications

Real-time relative qPCR without reference to control samples and estimation of run-specific PCR parameters from run-internal mini-standard curves.

Similar Datasets

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