tests/testthat/test_dispersions.R

context("dispersions")
test_that("expected errors thrown during dispersion estimation", {
  dds <- makeExampleDESeqDataSet(n=100, m=2)
  dds <- estimateSizeFactors(dds)
  expect_error(estimateDispersionsGeneEst(dds))

  set.seed(1)
  dds <- makeExampleDESeqDataSet(n=100, m=4, dispMeanRel=function(x) 0.001 + x/1e3, interceptMean=8, interceptSD=2)
  dds <- estimateSizeFactors(dds)
  mcols(dds)$dispGeneEst <- rep(1e-7, 100)
  expect_error(estimateDispersionsFit(dds))
  dds <- estimateDispersionsGeneEst(dds)
  expect_message(estimateDispersionsFit(dds))

  dds <- makeExampleDESeqDataSet(n=100, m=4)
  dds <- estimateSizeFactors(dds)
  mcols(dds)$dispGeneEst <- rep(1e-7, 100)
  dispersionFunction(dds) <- function(x) 1e-6
  expect_warning(estimateDispersionsMAP(dds))

  dds <- makeExampleDESeqDataSet(n=100, m=4)
  dds <- estimateSizeFactors(dds)
  levels(dds$condition) <- c("A","B","C")
  expect_error(estimateDispersions(dds))
  dds$condition <- droplevels(dds$condition)
  dds$group <- dds$condition
  design(dds) <- ~ group + condition
  expect_error(estimateDispersions(dds))

  dds <- makeExampleDESeqDataSet(n=100, m=2)
  expect_error({ dds <- DESeq(dds) })
  
})

test_that("the fitting of dispersion gives expected values using various methods", {
  # test the optimization of the logarithm of dispersion (alpha)
  # parameter with Cox-Reid adjustment and prior distribution.
  # also test the derivatives of the log posterior w.r.t. log alpha
  m <- 10
  set.seed(1)
  y <- rpois(m,20)
  sf <- rep(1,m)
  condition <- factor(rep(0:1,each=m/2))
  x <- cbind(rep(1,m),rep(0:1,each=m/2))
  colnames(x) <- c("Intercept","condition")

  lambda <- 2
  alpha <- .5

  # make a DESeqDataSet but don't use the design formula
  # instead we supply a model matrix below
  dds <- DESeqDataSetFromMatrix(matrix(y,nrow=1),
                                colData=DataFrame(condition),
                                design= ~ condition)
  sizeFactors(dds) <- sf
  dispersions(dds) <- alpha
  mcols(dds)$baseMean <- mean(y)

  # for testing we convert beta to the naturual log scale:
  # convert lambda from log to log2 scale by multiplying by log(2)^2
  # then convert beta back from log2 to log scale by multiplying by log(2)
  betaDESeq <- log(2)*DESeq2:::fitNbinomGLMs(dds, lambda=c(0,lambda*log(2)^2),modelMatrix=x)$betaMatrix
  log_alpha_prior_mean <- .5
  log_alpha_prior_sigmasq <- 1
  mu.hat <- as.numeric(exp(x %*% t(betaDESeq)))
  
  dispRes <- DESeq2:::fitDisp(ySEXP = matrix(y,nrow=1), xSEXP = x,
                              mu_hatSEXP = matrix(mu.hat,nrow=1), log_alphaSEXP = 0,
                              log_alpha_prior_meanSEXP = log_alpha_prior_mean,
                              log_alpha_prior_sigmasqSEXP = log_alpha_prior_sigmasq,
                              min_log_alphaSEXP = log(1e-8), kappa_0SEXP = 1,
                              tolSEXP = 1e-16, maxitSEXP = 100, usePriorSEXP = TRUE,
                              weightsSEXP=matrix(1,nrow=1,ncol=length(y)),
                              useWeightsSEXP=FALSE,
                              weightThresholdSEXP=1e-2,
                              useCRSEXP=TRUE)
  
  # maximum a posteriori (MAP) estimate from DESeq
  dispDESeq <- dispRes$log_alpha
  
  # MAP estimate using optim
  logPost <- function(log.alpha) {
    alpha <- exp(log.alpha)
    w <- diag(1/(1/mu.hat^2 * ( mu.hat + alpha * mu.hat^2 )))
    logLike <- sum(dnbinom(y, mu=mu.hat, size=1/alpha, log=TRUE))
    coxReid <- -.5*(log(det(t(x) %*% w %*% x)))
    logPrior <- dnorm(log.alpha, log_alpha_prior_mean, sqrt(log_alpha_prior_sigmasq), log=TRUE)
    (logLike + coxReid + logPrior)
  }
  
  dispOptim <- optim(0, function(p) -1*logPost(p), control=list(reltol=1e-16), method="Brent", lower=-10, upper=10)$par
                     
  expect_equal(dispDESeq, dispOptim, tolerance=1e-6)
  
  # check derivatives:
  
  # from Ted Harding https://stat.ethz.ch/pipermail/r-help/2007-September/140013.html
  num.deriv <- function(f,x,h=0.001) (f(x + h/2) - f(x-h/2))/h
  num.2nd.deriv <- function(f,x,h=0.001) (f(x + h) - 2*f(x) + f(x - h))/h^2

  # first derivative of log posterior w.r.t log alpha at start
  dispDerivDESeq <- dispRes$initial_dlp
  dispDerivNum <- num.deriv(logPost,0)

  expect_equal(dispDerivDESeq, dispDerivNum, tolerance=1e-6)

  # second derivative at finish
  dispD2DESeq <- dispRes$last_d2lp
  dispD2Num <- num.2nd.deriv(logPost, dispRes$log_alpha)

  expect_equal(dispD2DESeq, dispD2Num, tolerance=1e-6)


  # test fit alternative
  dds <- makeExampleDESeqDataSet()
  dds <- estimateSizeFactors(dds)
  ddsLocal <- estimateDispersions(dds, fitType="local")
  ddsMean <- estimateDispersions(dds, fitType="mean")
  ddsMed <- estimateDispersionsGeneEst(dds)
  useForMedian <- mcols(ddsMed)$dispGeneEst > 1e-7
  medianDisp <- median(mcols(ddsMed)$dispGeneEst[useForMedian],na.rm=TRUE)
  dispersionFunction(ddsMed) <- function(mu) medianDisp
  ddsMed <- estimateDispersionsMAP(ddsMed)  


  # test iterative
  set.seed(1)
  dds <- makeExampleDESeqDataSet(m=50,n=100,betaSD=1,interceptMean=8)
  dds <- estimateSizeFactors(dds)
  dds <- estimateDispersionsGeneEst(dds, niter=5)
  with(mcols(dds)[!mcols(dds)$allZero,],
       expect_equal(log(trueDisp), log(dispGeneEst),tol=0.2))

})

## test_that("Expected variance of log dispersions for df <= 3", {
##   sds <- rep(seq(from=.5, to=2.5, by=.25), each=2)
##   ests <- numeric(length(sds))
##   for (i in seq_along(sds)) {
##     cat(i)
##     dds <- makeExampleDESeqDataSet(n=1000, m=4, interceptMean=8, interceptSD=1,
##                                    dispMeanRel=function(x) exp(rnorm(1000,log(0.05),sds[i])))
##     sizeFactors(dds) <- rep(1,4)
##     dds <- estimateDispersions(dds, fitType="mean", quiet=TRUE)
##     ests[i] <- attr(dispersionFunction(dds), "dispPriorVar")
##   }
##   plot(sds^2, ests); abline(0,1)
## })

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DESeq2 documentation built on Feb. 22, 2021, 10 a.m.