Real-Time Quantitative PCR - Primer Design, Reference Gene Selection, Calculations and Statistics

A - Papers appearing in refereed journals

Chen, Z., Halford, N. G. and Liu, C. 2023. Real-Time Quantitative PCR - Primer Design, Reference Gene Selection, Calculations and Statistics. Metabolites. 13 (7), p. 806.

AuthorsChen, Z., Halford, N. G. and Liu, C.

Real-time quantitative PCR is a technique that can measure the content of the target nucleic acid sequence of interest in a given sample. It is mainly divided into absolute and relative quantitative methods. The relative quantification is mainly used in gene expressions for functional genomic and
transcriptome studies. However, to use this technology accurately, there are some key points to master. First, specific primers need to be designed to ensure amplification of the gene of interest (GOI). Second, the appropriate reference gene or reference gene combination has to be selected. Finally, scientific gene expression level calculations and statistics are required to obtain accurate
results. Therefore, this work proposes a workflow for relative quantitative PCR and introduces the relevant points so that beginners can better understand and use this technology.

KeywordsQuantitative real-time PCR; Relative gene expression; Normalized relative quantity; PCR efficiency
Year of Publication2023
Journal citation13 (7), p. 806
Digital Object Identifier (DOI)
Web address (URL)
Open accessPublished as ‘gold’ (paid) open access
FunderBiotechnology and Biological Sciences Research Council
Funder project or codeDesigning Future Wheat (DFW) [ISPG]
Publisher's version
Output statusPublished
Publication dates
Online28 Jun 2023
Publication process dates
Accepted27 Jun 2023

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