Description Usage Arguments Details Value Author(s) References See Also Examples
Given a microarray linear model fit, compute moderated t-statistics, moderated F-statistic, and log-odds of differential expression by empirical Bayes moderation of the standard errors towards a common value.
1 2 3 |
fit |
an |
proportion |
numeric value between 0 and 1, assumed proportion of genes which are differentially expressed |
stdev.coef.lim |
numeric vector of length 2, assumed lower and upper limits for the standard deviation of log2-fold-changes for differentially expressed genes |
trend |
logical, should an intensity-trend be allowed for the prior variance? Default is that the prior variance is constant. |
robust |
logical, should the estimation of |
winsor.tail.p |
numeric vector of length 1 or 2, giving left and right tail proportions of |
lfc |
the minimum log2-fold-change that is considered scientifically meaningful |
These functions is used to rank genes in order of evidence for differential expression.
They use an empirical Bayes method to shrink the probe-wise sample variances towards a common value and to augmenting the degrees of freedom for the individual variances (Smyth, 2004).
The functions accept as input argument fit
a fitted model object from the functions lmFit
, lm.series
, mrlm
or gls.series
.
The fitted model object may have been processed by contrasts.fit
before being passed to eBayes
to convert the coefficients of the design matrix into an arbitrary number of contrasts which are to be tested equal to zero.
The columns of fit
define a set of contrasts which are to be tested equal to zero.
The empirical Bayes moderated t-statistics test each individual contrast equal to zero. For each probe (row), the moderated F-statistic tests whether all the contrasts are zero. The F-statistic is an overall test computed from the set of t-statistics for that probe. This is exactly analogous the relationship between t-tests and F-statistics in conventional anova, except that the residual mean squares and residual degrees of freedom have been moderated between probes.
The estimates s2.prior
and df.prior
are computed by fitFDist
.
s2.post
is the weighted average of s2.prior
and sigma^2
with weights proportional to df.prior
and df.residual
respectively.
The lods
is sometimes known as the B-statistic.
The F-statistics F
are computed by classifyTestsF
with fstat.only=TRUE
.
eBayes
doesn't compute ordinary (unmoderated) t-statistics by default, but these can be easily extracted from
the linear model output, see the example below.
ebayes
is the earlier and leaner function, kept for background capatability, while
eBayes
is the later more object-orientated version.
The difference is that ebayes
outputs only the empirical Bayes statistics whereas eBayes
adds them to the fitted model object fit
.
eBayes
is recommended for routine use as it produces objects containing all the necessary components for downstream analysis
treat
computes empirical Bayes moderated-t p-values relative to a minimum required fold-change threshold.
Use topTreat
to summarize output from treat
.
Instead of testing for genes which have log-fold-changes different from zero, it tests whether the log2-fold-change is greater than lfc
in absolute value (McCarthy and Smyth, 2009).
treat
is concerned with p-values rather than posterior odds, so it does not compute the B-statistic lods
.
The idea of thresholding doesn't apply to F-statistics in a straightforward way, so moderated F-statistics are also not computed.
If trend=TRUE
then an intensity-dependent trend is fitted to the prior variances s2.prior
.
Specifically, squeezeVar
is called with the covariate
equal to Amean
, the average log2-intensity for each gene.
See squeezeVar
for more details.
If robust=TRUE
then the robust empirical Bayes procedure of Phipson et al (2013) is used.
See squeezeVar
for more details.
eBayes
produces an object of class MArrayLM
(see MArrayLM-class
) containing everything found in fit
plus the following added components:
t |
numeric vector or matrix of moderated t-statistics |
p.value |
numeric vector of p-values corresponding to the t-statistics |
s2.prior |
estimated prior value for |
df.prior |
degrees of freedom associated with |
df.total |
numeric vector of total degrees of freedom associated with t-statistics and p-values. Equal to |
s2.post |
numeric vector giving the posterior values for |
lods |
numeric vector or matrix giving the log-odds of differential expression |
var.prior |
estimated prior value for the variance of the log2-fold-change for differentially expressed gene |
F |
numeric vector of moderated F-statistics for testing all contrasts defined by the columns of |
F.p.value |
numeric vector giving p-values corresponding to |
treat
a produces an MArrayLM
object similar to eBayes
but without lods
, var.prior
, F
or F.p.value
.
ebayes
produces an ordinary list containing the above components except for F
and F.p.value
.
Gordon Smyth and Davis McCarthy
McCarthy, D. J., and Smyth, G. K. (2009). Testing significance relative to a fold-change threshold is a TREAT. Bioinformatics. http://bioinformatics.oxfordjournals.org/cgi/content/abstract/btp053
Loennstedt, I., and Speed, T. P. (2002). Replicated microarray data. Statistica Sinica 12, 31-46.
Phipson, B, Lee, S, Majewski, IJ, Alexander, WS, and Smyth, GK (2013). Empirical Bayes in the presence of exceptional cases, with application to microarray data. Bioinformatics Division, Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia. http://www.statsci.org/smyth/pubs/RobustEBayesPreprint.pdf
Smyth, G. K. (2004). Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Statistical Applications in Genetics and Molecular Biology, Volume 3, Article 3. http://www.statsci.org/smyth/pubs/ebayes.pdf
squeezeVar
, fitFDist
, tmixture.matrix
.
An overview of linear model functions in limma is given by 06.LinearModels.
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 | # See also lmFit examples
# Simulate gene expression data,
# 6 microarrays and 100 genes with one gene differentially expressed
set.seed(2004); invisible(runif(100))
M <- matrix(rnorm(100*6,sd=0.3),100,6)
M[1,] <- M[1,] + 1
fit <- lmFit(M)
# Moderated t-statistic
fit <- eBayes(fit)
topTable(fit)
# Ordinary t-statistic
ordinary.t <- fit$coef / fit$stdev.unscaled / fit$sigma
# Q-Q plots of t statistics
# Points off the line may be differentially expressed
par(mfrow=c(1,2))
qqt(ordinary.t, df=fit$df.residual, main="Ordinary t")
abline(0,1)
qqt(fit$t, df=fit$df.total,main="Moderated t")
abline(0,1)
par(mfrow=c(1,1))
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