To cite freqpcr in publications use:
Sudo M, Osakabe M (2021). “freqpcr: Estimation of Population Allele Frequency Using qPCR ΔΔCq Measures From Bulk Samples.” Molecular Ecology Resources, n/a(n/a), 0–14. doi:10.1111/1755-0998.13554, https://onlinelibrary.wiley.com/doi/pdf/10.1111/1755-0998.13554, https://onlinelibrary.wiley.com/doi/abs/10.1111/1755-0998.13554.
Corresponding BibTeX entry:
@Article{,
title = {freqpcr: Estimation of Population Allele Frequency Using
qPCR ΔΔCq Measures From Bulk Samples},
author = {Masaaki Sudo and Masahiro Osakabe},
journal = {Molecular Ecology Resources},
year = {2021},
volume = {n/a},
number = {n/a},
pages = {0--14},
keywords = {confidence interval, group testing, maximum-likelihood
estimation, R language, real-time polymerase chain reaction},
doi = {10.1111/1755-0998.13554},
url =
{https://onlinelibrary.wiley.com/doi/abs/10.1111/1755-0998.13554},
eprint =
{https://onlinelibrary.wiley.com/doi/pdf/10.1111/1755-0998.13554},
abstract = {PCR techniques, both quantitative (qPCR) and
nonquantitative, have been used to estimate the frequency of a
specific allele in a population. However, the labour required to
sample numerous individuals and subsequently handle each sample
renders the quantification of rare mutations (e.g., pesticide
resistance gene mutations at the early stages of resistance
development) challenging. Meanwhile, pooling DNA from multiple
individuals as a “bulk sample” combined with qPCR may reduce
handling costs. The qPCR output for a bulk sample, however,
contains uncertainty owing to variations in DNA yields from each
individual, in addition to measurement errors. In this study, we
have developed a statistical model to estimate the frequency of
the specific allele and its confidence interval when the sample
allele frequencies are obtained in the form of ΔΔCq in the qPCR
analyses on multiple bulk samples collected from a population. We
assumed a gamma distribution as the individual DNA yield and
developed an R package for parameter estimation, which was
verified using real DNA samples from acaricide-resistant spider
mites, as well as a numerical simulation. Our model resulted in
unbiased point estimates of the allele frequency compared with
simple averaging of the ΔΔCq values. The confidence intervals
suggest that dividing the bulk samples into more parts will
improve precision if the total number of individuals is equal;
however, if the cost of PCR analysis is higher than that of
sampling, increasing the total number and pooling them into a few
bulk samples may also yield comparable precision.},
}