ttestCtData: Differentially expressed features with qPCR: t-test

In HTqPCR: Automated analysis of high-throughput qPCR data

Description

Function for calculating t-test and p-values across two groups for the features present in high-throughput qPCR data, such as from TaqMan Low Density Arrays.

Usage

ttestCtData(q, groups = NULL, calibrator, alternative = "two.sided", paired = FALSE, replicates = TRUE, sort = TRUE, stringent = TRUE, p.adjust = "BH", ...)

Arguments

q	qPCRset object.
groups	factor, assigning each sample to one of two groups.
calibrator	which of the two groups is to be considered as the reference and not the test? Defaults to the first group in groups.
alternative	character string (first letter is enough), specifying the alternative hypothesis, "two.sided" (default), "greater" or "less".
paired	logical, should a paired t-test be used.
replicates	logical, if replicated genes are present on the array, the statistics will be calculated for all the replicates combined, rather than the individual wells.
sort	boolean, should the output be sorted by p-values.
stringent	boolean, for flagging results as "Undetermined". See details.
p.adjust	character string, which method to use for p-value adjustment for multiple testing. See details.
...	any other arguments will be passed to the t.test function.

Details

Once the Ct values have been normalised, differential expression can be calculated. This function deals with just the simple case, where there are two types of samples to compare. For more complex studies, see limmaCtData.

All results are assigned to a category, either "OK" or "Undetermined" depending on the input Ct values. If stringent=TRUE any unreliable or undetermined measurements among technical and biological replicates will result in the final result being "Undetermined". For stringent=FALSE the result will be "OK" unless at least half of the Ct values for a given gene are unreliable/undetermined.

The argument p.adjust is passed on to the p.adjust function. Options include e.g. "BH" (Benjamini & Hochberg, the default), "fdr" and "bonferroni". See p.adjust for more information on the individual methods.

Value

A data.frame containing the following information:

genes	The names of the features on the card.
feature.pos	The featurePos of the genes. If replicated genes are used, the feature positions will be concatenated together.
t.test	The value of the t-test.
p.value	The corresponding p-value.
ddCt	The delta delta Ct values.
FC	The fold change; 2^(-ddCt).
meanCalibrator	The average expression level of each gene in the calibrator sample(s).
meanTarget	The average expression level of each gene in the target sample(s).
categoryCalibrator	The category of the Ct values ("OK", "Undetermined") across the calibrator.
categoryTarget	Ditto for the target.

Author(s)

Heidi Dvinge

See Also

t.test, limmaCtData, mannwhitneyCtData. plotCtRQ and plotCtSignificance can be used for visualising the results.

HTqPCR documentation built on Nov. 8, 2020, 6:51 p.m.