varFit(data, design = NULL, coef = NULL, type = NULL, trend = TRUE, robust = TRUE, weights = NULL)MethylSet or matrix with rows corresponding to the features of interest such as CpG sites and columns corresponding to samples or arrays"AD" for absolute residuals or "SQ" for squared residuals. Default is absolute.MArrayLM (see MArrayLM-class) containing everything found in a fitted model object produced by lmFit and eBayes as well as a vector containing the sample CpG-wise variances and a matrix of LogVarRatios corresponding to the differential variability analysis.limma package and is used to rank features such as CpG sites or genes in order of evidence of differential variability between different comparisons corresponding to the columns of the design matrix. A measure of variability is calculated for each CpG in each sample by subtracting out the group mean and taking the absolute or squared deviation. A linear model is then fitted to the absolute or squared deviations. The residuals of the linear model fit are subjected to empirical Bayes shrinkage and moderated t statistics (Smyth, 2004) calculated. False discovery rates are calculated using the method of Benjamini and Hochberg (1995).If coef is not specified, then group means are estimated based on all the columns of the design matrix and subtracted out before testing for differential variability. If the design matrix contains nuisance parameters, then subsetting the design matrix columns by coef should remove these columns from the design matrix. If the design matrix includes an intercept term, this should be included in coef. The nuisance parameters are included in the linear model fit to the absolute or squared deviations, but should not be considered when subtracting group means to obtain the deviations. Note that design matrices without an intercept term are permitted, and specific contrasts tested using the function contrasts.varFit.
For methylation data, the analysis is performed on the M-values, defined as the log base 2 ratio of the methylated signal to the unmethylated signal. If a MethylSet object is supplied, M-values are extracted with an offset of 100 added to the numerator and denominator.
For testing differential variability on RNA-Seq data, a DGEList object can be supplied directly to the function. A voom transformation is applied before testing for differential variability. The weights calculated in voom are used in the linear model fit.
Since the output is of class MArrayLM, any functions that can be applied to fit objects from lmFit and eBayes can be applied, for example, topTable and decideTests.
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.
Smyth, G. K. (2005). Limma: linear models for microarray data. In: Bioinformatics and Computational Biology Solutions using R and Bioconductor. R. Gentleman, V. Carey, S. Dudoit, R. Irizarry, W. Huber (eds), Springer, New York, 2005.
Benjamini, Y., and Hochberg, Y. (1995). Controlling the false discovery rate: a practical and powerful approach to multiple testing. Journal of the Royal Statistical Society Series, B, 57, 289-300.
contrasts.varFit, topVar, getLeveneResiduals, lmFit, eBayes, topTable, decideTests, voom# Randomly generate data for a 2 group problem with 100 CpG sites and 5 arrays in each group.
y<-matrix(rnorm(1000),ncol=10)
group<-factor(rep(c(1,2),each=5))
design<-model.matrix(~group)
# Fit linear model for differential variability
vfit<-varFit(y,design,coef=c(1,2))
# Look at top table of results
topVar(vfit,coef=2)Run the code above in your browser using DataLab