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Real-Time PCR for mRNA Quantitation

Open access
Wong, Marisa L., Medrano, Juan F.·2005·BioTechniques
DOI 10.2144/05391rv01
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Abstract

Real-time PCR has become one of the most widely used methods of gene quantitation because it has a large dynamic range, boasts tremendous sensitivity, can be highly sequence-specific, has little to no post-amplification processing, and is amenable to increasing sample throughput. However, optimal benefit from these advantages requires a clear understanding of the many options available for running a real-time PCR experiment. Starting with the theory behind real-time PCR, this review discusses the key components of a real-time PCR experiment, including one-step or two-step PCR, absolute versus relative quantitation, mathematical models available for relative quantitation and amplification efficiency calculations, types of normalization or data correction, and detection chemistries. In addition, the many causes of variation as well as methods to calculate intra- and inter-assay variation are addressed.

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Metadata

Authors
Wong, M. L., & Medrano, J. F.
Venue
BioTechniques
Publisher
Informa UK Limited
Published
2005-07
Volume
39
Issue
1
Pages
75-85
ISSN
  • 0736-6205 (ISSN-L)
  • 1940-9818
DOI
10.2144/05391rv01
OpenAlex
W2122749842
Cited by
1,786
References
1
FWCI
27.08
Type
journal article
Language
en
Open access
gold
Licence
creativecommons.org/licenses/by/4.0/ (vor)
Funders
  • National Institutes of Health (NIH T32-GM08799-01)
  • U.S. Department of Agriculture (2001-52100-11211)

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Wong, M. L., & Medrano, J. F. (2005). Real-Time PCR for mRNA Quantitation. BioTechniques. https://doi.org/10.2144/05391rv01

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