Abstract:
BACKGROUND:A recently proposed method for estimating qPCR amplification efficiency E analyzes fluorescence intensity ratios from pairs of points deemed to lie in the exponential growth region on the amplification curves for all reactions in a dilution series. This method suffers from a serious problem: The resulting ratios are highly correlated, as they involve multiple use of the raw data, for example, yielding ~ 250 E estimates from ~ 25 intensity readings. The resulting statistics for such estimates are falsely optimistic in their assessment of the estimation precision. RESULTS:Monte Carlo simulations confirm that the correlated pairs method yields precision estimates that are better than actual by a factor of two or more. This result is further supported by estimating E by both pairwise and Cq calibration methods for the 16 replicate datasets from the critiqued work, and then comparing the ensemble statistics for these methods. CONCLUSION:Contrary to assertions in the proposing work, the pairwise method does not yield E estimates a factor of 2 more precise than estimates from Cq calibration fitting (the standard curve method). On the other hand, the statistically correct direct fit of the data to the model behind the pairwise method can yield E estimates of comparable precision. Ways in which the approach might be improved are discussed briefly.
journal_name
BMC Bioinformaticsjournal_title
BMC bioinformaticsauthors
Tellinghuisen Jdoi
10.1186/s12859-020-03604-4subject
Has Abstractpub_date
2020-07-08 00:00:00pages
291issue
1issn
1471-2105pii
10.1186/s12859-020-03604-4journal_volume
21pub_type
杂志文章abstract:BACKGROUND:Existing large-scale metabolic models of sequenced organisms commonly include enzymatic functions which can not be attributed to any gene in that organism. Existing computational strategies for identifying such missing genes rely primarily on sequence homology to known enzyme-encoding genes. RESULTS:We pres...
journal_title:BMC bioinformatics
pub_type: 杂志文章
doi:10.1186/1471-2105-7-177
更新日期:2006-03-29 00:00:00
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journal_title:BMC bioinformatics
pub_type: 杂志文章
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更新日期:2018-09-26 00:00:00
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