| Title: | Call wide peaks for sequencing data |
|---|---|
| Description: | Nucleolus is an important structure inside the nucleus in eukaryotic cells. It is the site for transcribing rDNA into rRNA and for assembling ribosomes, aka ribosome biogenesis. In addition, nucleoli are dynamic hubs through which numerous proteins shuttle and contact specific non-rDNA genomic loci. Deep sequencing analyses of DNA associated with isolated nucleoli (NAD- seq) have shown that specific loci, termed nucleolus- associated domains (NADs) form frequent three- dimensional associations with nucleoli. NAD-seq has been used to study the biological functions of NAD and the dynamics of NAD distribution during embryonic stem cell (ESC) differentiation. Here, we developed a Bioconductor package NADfinder for bioinformatic analysis of the NAD-seq data, including baseline correction, smoothing, normalization, peak calling, and annotation. |
| Authors: | Jianhong Ou, Haibo Liu, Jun Yu, Hervé Pagès, Paul Kaufman, Lihua Julie Zhu |
| Maintainer: | Jianhong Ou <[email protected]>, Lihua Julie Zhu <[email protected]> |
| License: | GPL (>= 2) |
| Version: | 1.31.0 |
| Built: | 2024-10-30 08:31:09 UTC |
| Source: | https://github.com/bioc/NADfinder |
Sliding-window based peak calling algorithm using whole genome sequences as control
Correct ratios of read counts per sliding window for background.
backgroundCorrection(ratios, degree = 3, ...)backgroundCorrection(ratios, degree = 3, ...)
ratios |
A vector of numeric. It is log2-transformed ratios, CPMRatios or OddRatios of counts for each window. |
degree |
Degree of polynomial. default 3. |
... |
parameters could be passed to baseline.modpolyfit. |
This function implements the backgound correction methods of
algorithm for polynomial fitting. See details via
baseline.modpolyfit. This function expects
the trendency of decreasing of the ratios from 5' end to 3' end.
A vector of numeric. It is the background corrected log2-transformed ratios, CPMRatios or OddRatios.
x <- runif(200) background <- rep(c(20:1)/100, each=10) backgroundCorrection(x)x <- runif(200) background <- rep(c(20:1)/100, each=10) backgroundCorrection(x)
The Butterworth filter is a type of signal processing filter designed to have as flat a frequency response as possible in the passband.
butterFilter(ratios, N = ceiling(length(ratios)/200))butterFilter(ratios, N = ceiling(length(ratios)/200))
ratios |
A vector of numeric. It is log2-transformed ratios, CPMRatios or OddRatios in each window. |
N |
numeric(1) or integer(1). Critical frequencies of the low pass filter will be set as 1/N. 1/N is a cutoff at 1/N-th of the Nyquist frequency. By default, it is suppose there are about 200 peaks in the inputs. |
A vector of numeric with same length of input ratios. The vector indicates smoothed ratios.
ratios <- runif(20000) butterFilter(ratios)ratios <- runif(20000) butterFilter(ratios)
Use limma to calculate p-values for NADs
callPeaks( se, backgroundCorrectedAssay = "bcRatio", normalization.method = "quantile", N = 100, cutoffAdjPvalue = 1e-04, countFilter = 1000, combineP.method = "minimump", smooth.method = "loess", lfc = log2(1.5), ... )callPeaks( se, backgroundCorrectedAssay = "bcRatio", normalization.method = "quantile", N = 100, cutoffAdjPvalue = 1e-04, countFilter = 1000, combineP.method = "minimump", smooth.method = "loess", lfc = log2(1.5), ... )
se |
An object of RangedSummarizedExperiment with assays of raw counts, tranformed ratios, background corrected ratios, smoothed ratios and z-scores. It should be an element of output of smoothRatiosByChromosome |
backgroundCorrectedAssay |
character(1). Assays names for background corrected log2-transformed ratios, CPMRatios or OddRatios. |
normalization.method |
character(1) specifying the normalization method to be used. Choices are "none", "scale", "quantile" or "cyclicloess". See normalizeBetweenArrays for details. |
N |
numeric(1) or integer(1). The number of neighboring windows used for loess smoothing or the inverse of the critical frequencies of the low pass filter for butterworth filter. 1/N is a cutoff at 1/N-th of the Nyquist frequency. Default 100. |
cutoffAdjPvalue |
numeric(1). Cutoff adjust p-value. |
countFilter |
numeric(1). Cutoff value for mean of raw reads count in each window. |
combineP.method |
A method used to combine P-values. Default minimump |
smooth.method |
A method used to smooth the ratios. Choices are "loess", "none" and "butterworthfilter". |
lfc |
the minimum log2-fold-change that is considered scientifically meaningful |
... |
Parameter not used. |
By default, use the mean smoothed ratio for each peak region to calculate p-values
An object of GRanges of peak list with metadata "AveSig", "P.Value", and "adj.P.Val", where "AveSig" means average signal such as average log2OddsRatio, log2CPMRatio or log2Ratio.
Jianhong Ou, Haibo Liu and Julie Zhu
data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) se<- smoothRatiosByChromosome(se, chr="chr18") #add some variability to the data since the triplicate.count data was created using one sample only assays(se[[1]])$bcRatio[,2] <- assays(se[[1]])$bcRatio[,2] + 0.3 assays(se[[1]])$bcRatio[,3] <- assays(se[[1]])$bcRatio[,3] - 0.3 peaks <- callPeaks(se[[1]], cutoffAdjPvalue=0.001, countFilter=10)data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) se<- smoothRatiosByChromosome(se, chr="chr18") #add some variability to the data since the triplicate.count data was created using one sample only assays(se[[1]])$bcRatio[,2] <- assays(se[[1]])$bcRatio[,2] + 0.3 assays(se[[1]])$bcRatio[,3] <- assays(se[[1]])$bcRatio[,3] - 0.3 peaks <- callPeaks(se[[1]], cutoffAdjPvalue=0.001, countFilter=10)
Perform overlap queries between reads and genome by sliding windows Count reads over sliding windows.
computeLibSizeChrom(aln_list)computeLibSizeChrom(aln_list)
aln_list |
a list. |
A RangedSummarizedExperiment object with chromosome-level depth The assays slot holds the counts, rowRanges holds the annotation from the sliding widows of genome. metadata contains lib.size.chrom for holding chromosome-level sequence depth
Jun Yu,Hervé Pagès and Julie Zhu
Plot the difference between the cumulative percentage of tag allocation in paired samples.
cumulativePercentage( se, binWidth = 1e+05, backgroundCorrectedAssay = "bcRatio", ... )cumulativePercentage( se, binWidth = 1e+05, backgroundCorrectedAssay = "bcRatio", ... )
se |
An object of RangedSummarizedExperiment with assays of raw counts, transfomred ratios, background correct ratios, smoothed ratios and z-scores. It should be an element of the output of smoothRatiosByChromosome. |
binWidth |
numeric(1) or integer(1). The width of each bin. |
backgroundCorrectedAssay |
character(1). Assays names for background correction ratios. |
... |
Parameter not used. |
A list of data.frame with the cumulative percentages.
Normalization, bias correction, and peak calling for ChIP-seq Aaron Diaz, Kiyoub Park, Daniel A. Lim, Jun S. Song Stat Appl Genet Mol Biol. Author manuscript; available in PMC 2012 May 3.Published in final edited form as: Stat Appl Genet Mol Biol. 2012 Mar 31; 11(3): 10.1515/1544-6115.1750 /j/sagmb.2012.11.issue-3/1544-6115.1750/1544-6115.1750.xml. Published online 2012 Mar 31. doi: 10.1515/1544-6115.1750 PMCID: PMC3342857
library(SummarizedExperiment) data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) se <- smoothRatiosByChromosome(se, chr="chr18") cumulativePercentage(se[["chr18"]])library(SummarizedExperiment) data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) se <- smoothRatiosByChromosome(se, chr="chr18") cumulativePercentage(se[["chr18"]])
Output signals to bedgraph, bed, wig, etc, for track viewer
exportSignals(dat, assayName, colName, con, format = "bedGraph", ...)exportSignals(dat, assayName, colName, con, format = "bedGraph", ...)
dat |
An object of GRanges, or RangedSummarizedExperiment with assays of raw counts, ratios, background correct ratios, smoothed ratios and z-scores. It should be an element of output of smoothRatiosByChromosome |
assayName |
character(1). Assay name for RangedSummarizedExperiment |
colName |
character(1). Column name of metadata of dat or assay of dat for coverage weight, see coverage, RangedSummarizedExperiment. |
con |
The connection to which data is saved. If this is a character vector, it is assumed to be a filename and a corresponding file connection is created and then closed after exporting the object. If missing, a SimpleRleList will be returned. |
format |
The format of the output. see export. |
... |
Parameters to be passed to export |
If con is missing, a SimpleRleList will be returned. Otherwise, nothing is returned.
gr <- GRanges("chr1", IRanges(seq_len(100), 201:300), reads=rep(1, 100)) myTrackLine <- new("TrackLine", name="my track", description="description of my track", color=col2rgb("red")[, 1], visibility="full") exportSignals(gr, colName="reads", con="test.bedGraph", trackLine=myTrackLine) data(triplicate.count) exportSignals(triplicate.count, "counts", "G18.subsampled.srt.bam", "test.bw", format="bigWig")gr <- GRanges("chr1", IRanges(seq_len(100), 201:300), reads=rep(1, 100)) myTrackLine <- new("TrackLine", name="my track", description="description of my track", color=col2rgb("red")[, 1], visibility="full") exportSignals(gr, colName="reads", con="test.bedGraph", trackLine=myTrackLine) data(triplicate.count) exportSignals(triplicate.count, "counts", "G18.subsampled.srt.bam", "test.bw", format="bigWig")
Get correlations and p-values between biological replicates based on coverage signal for peak regions. The signals will be filtered by the background cutoff value before calculated correlations. This function also output a correlation plots using the corrplot.
getCorrelations( se, chr = paste0("chr", seq_len(19)), ratioAssay = "ratio", window = 10000L, cutoff = 1, method = c("spearman", "pearson", "kendall"), file_name = "Correlation plots.pdf", ... )getCorrelations( se, chr = paste0("chr", seq_len(19)), ratioAssay = "ratio", window = 10000L, cutoff = 1, method = c("spearman", "pearson", "kendall"), file_name = "Correlation plots.pdf", ... )
se |
A RangedSummarizedExperiment object. The output from log2se. |
chr |
A vector of character. Filter for seqnames. It should be the chromosome names to be kept. |
ratioAssay |
character(1). Column name of ratio for correlation calculation. |
window |
numeric(1) or integer(1). The window size for summary of the ratios. |
cutoff |
numeric(1). All the coverage signals lower than cutoff value in a given window will be filtered out. |
method |
character(1) indicating which correlation coefficient is to be computed. See cor. |
file_name |
A file name for output correlation plots |
... |
Parameters not used. |
A list of matrixes of correlation coefficients and p-values.
Jianhong Ou, Haibo Liu
data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) getCorrelations(se, chr="chr18")data(triplicate.count) se <- triplicate.count se <- log2se(se, transformation = "log2CPMRatio", nucleolusCols = c("N18.subsampled.srt-2.bam", "N18.subsampled.srt-3.bam", "N18.subsampled.srt.bam"), genomeCols = c("G18.subsampled.srt-2.bam", "G18.subsampled.srt-3.bam", "G18.subsampled.srt.bam")) getCorrelations(se, chr="chr18")
Detect peaks and calculate z-scores for each peak
groupZscores(zscore)groupZscores(zscore)
zscore |
A vector of numeric. It is the z-scores of ratios for each window. |
A data.frame with column names as "zscore", "group", "grp.zscore", and "pvalue".
x <- seq_len(500) a <- 2 * 2*pi/length(x) y <- 20 * sin(x*a) noise1 <- 20 * 1/10 * sin(x*a*10) zscore <- y+noise1 groupZscores(zscore)x <- seq_len(500) a <- 2 * 2*pi/length(x) y <- 20 * sin(x*a) noise1 <- 20 * 1/10 * sin(x*a*10) zscore <- y+noise1 groupZscores(zscore)
Count reads overlapping a set of genimc features represented as genomic ranges. This function does not work for parallel.
IntersectionNotStrict( features, reads, ignore.strand = TRUE, inter.feature = FALSE )IntersectionNotStrict( features, reads, ignore.strand = TRUE, inter.feature = FALSE )
features |
A object of GRanges representing the feature regions to be counted. |
reads |
An object that represents the data to be counted. See summarizeOverlaps. If reads are more than 1 bam files, it should be a vector of character with full path, otherwise current working directory is the default directory. For paired end reads, |
ignore.strand |
logical(1). ignore strand? |
inter.feature |
not used. This parameter is required by summarizeOverlaps. |
return a summarized experiment object with chromosome-level depth information for each input sample as metadata.
Calculate the log2 transformed ratios for nucleolus vs genome. pseudo-count will be used to avoid x/0 or log(0).
log2se( se, nucleolusCols, genomeCols, pseudocount = 1L, transformation = c("log2OddsRatio", "log2CPMRatio", "log2Ratio"), chrom.level.lib = TRUE )log2se( se, nucleolusCols, genomeCols, pseudocount = 1L, transformation = c("log2OddsRatio", "log2CPMRatio", "log2Ratio"), chrom.level.lib = TRUE )
se |
A RangedSummarizedExperiment object. The output of tileCount. |
nucleolusCols, genomeCols
|
column Names of counts for nucleolus and genome. They should be the column names in the assays of se. Ratios will be calculated as log2(transformed nucleolusCols/transformed genomeCols). |
pseudocount |
default to 1, pseudo-count used to aviod x/0 or log(0). |
transformation |
transformation type |
chrom.level.lib |
indicating whether calculating CPM or odds using sequence depth of the whole genome or the corresponding chromosome |
A RangedSummarizedExperiment object with log2 transformed ratios. Assays will be named as nucleolus, genome and ratio.
Jianhong Ou and Julie Zhu
library(SummarizedExperiment) se <- SummarizedExperiment(assays=list(counts=DataFrame(A=seq_len(3), B=rep(1, 3), C=rep(4, 3), D=rep(2, 3))), rowRanges=GRanges(c("chr1","chr1", "chr2"), IRanges(c(1, 10, 20), width=9))) metadata(se)$lib.size.chrom <- data.frame( c(1000, 1000), c(2000, 2000), c(200,200), c(300,300)) colnames(metadata(se)$lib.size.chrom) <- c("A", "B", "C", "D") rownames(metadata(se)$lib.size.chrom) <- c("chr1", "chr2") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2Ratio") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2CPMRatio") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2OddsRatio")library(SummarizedExperiment) se <- SummarizedExperiment(assays=list(counts=DataFrame(A=seq_len(3), B=rep(1, 3), C=rep(4, 3), D=rep(2, 3))), rowRanges=GRanges(c("chr1","chr1", "chr2"), IRanges(c(1, 10, 20), width=9))) metadata(se)$lib.size.chrom <- data.frame( c(1000, 1000), c(2000, 2000), c(200,200), c(300,300)) colnames(metadata(se)$lib.size.chrom) <- c("A", "B", "C", "D") rownames(metadata(se)$lib.size.chrom) <- c("chr1", "chr2") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2Ratio") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2CPMRatio") log2se(se, nucleolusCols = c("A", "C"), genomeCols = c("B", "D"), transformation = "log2OddsRatio")
Detect the peak positions and valley positions leveraging github::dgromer/peakdet
peakdet(y, delta = 0, silence = TRUE)peakdet(y, delta = 0, silence = TRUE)
y |
A numeric vector for searching peaks |
delta |
A numeric vector of length 1, defining the minimum absolute changes required for local maximum or minimum detection when slope sign changes. If it is set to 0, the delta will be set to 1/10 of the range of y. |
silence |
logical(1). If false, echo the delta value when delta is set as 0. |
A list with peakpos and valleypos. Both peakpos and valleypos are numeric vectors storing the positions of peaks or valleys.
y <- runif(200) peakdet(y) y <- sin(seq(0,20)) peakdet(y)y <- runif(200) peakdet(y) y <- sin(seq(0,20)) peakdet(y)
Plot signals with ideograms for GRangesList.
plotSig(ideo, grList, mcolName, ...)plotSig(ideo, grList, mcolName, ...)
ideo |
Output of loadIdeogram. |
grList |
A GRangesList of data to plot. |
mcolName |
Column name of metadata of GRangesList for plotting. |
... |
Parameters to pass to ideogramPlot |
Invisible argument list for ideogramPlot.
library(trackViewer) #ideo <- loadIdeogram("mm10") ideo <- readRDS(system.file("extdata", "ideo.mm10.rds", package = "NADfinder")) gr1 <- gr2 <- ideo mcols(gr1) <- DataFrame(score=runif(length(gr1))) mcols(gr2) <- DataFrame(score=runif(length(gr2))) grList <- GRangesList(gr1, gr2) plotSig(ideo, grList, mcolName="score", layout=list("chr1"))library(trackViewer) #ideo <- loadIdeogram("mm10") ideo <- readRDS(system.file("extdata", "ideo.mm10.rds", package = "NADfinder")) gr1 <- gr2 <- ideo mcols(gr1) <- DataFrame(score=runif(length(gr1))) mcols(gr2) <- DataFrame(score=runif(length(gr2))) grList <- GRangesList(gr1, gr2) plotSig(ideo, grList, mcolName="score", layout=list("chr1"))
Counts data for chromosome 18 for an experiment of a single pair of samples
Split the ratios by chromosome and do background correction and signal smoothing.
smoothRatiosByChromosome( se, chr = paste0("chr", c(seq_len(21), "X", "Y")), ratioAssay = "ratio", backgroundCorrectedAssay = "bcRatio", smoothedRatioAssay = "smoothedRatio", zscoreAssay = "zscore", backgroundPercentage = 0.25, chrom.level.background = TRUE, ... )smoothRatiosByChromosome( se, chr = paste0("chr", c(seq_len(21), "X", "Y")), ratioAssay = "ratio", backgroundCorrectedAssay = "bcRatio", smoothedRatioAssay = "smoothedRatio", zscoreAssay = "zscore", backgroundPercentage = 0.25, chrom.level.background = TRUE, ... )
se |
An object of RangedSummarizedExperiment with log2-transformed ratios, CPMRatios or OddRatios. Output of log2se |
chr |
A character vector, used to filter out seqnames. It should be the chromosome names to be kept. |
ratioAssay |
The name of assay in se, which store the values (log2-transformed ratios, CPMRatios or OddRatios) to be smoothed. |
backgroundCorrectedAssay, smoothedRatioAssay, zscoreAssay
|
character(1). Assays names for background corrected ratios, smoothed ratios and z-scores based on background corrected ratios. |
backgroundPercentage |
numeric(1). Percentage of values for background, see zscoreOverBck. The percentage of values lower than this threshold will be treated as background, with 25 percentile as default. |
chrom.level.background |
logical(1): TRUE or FALSE, default to TRUE, use chromosome-level background to calculate z-score |
... |
Parameters could be passed to butterFilter. |
A SimpleList of RangedSummarizedExperiment with smoothed ratios.
Jianhong Ou, Haibo Liu and Julie Zhu
data(single.count) se <- single.count dat <- log2se(se, nucleolusCols="N18.subsampled.srt.bam", genomeCols="G18.subsampled.srt.bam", transformation="log2CPMRatio") dat1 <- smoothRatiosByChromosome(dat, N=100, chr = c("chr18", "chr19")) dat2 <- smoothRatiosByChromosome(dat, N=100, chr = c("chr18", "chr19"), chrom.level.background = FALSE)data(single.count) se <- single.count dat <- log2se(se, nucleolusCols="N18.subsampled.srt.bam", genomeCols="G18.subsampled.srt.bam", transformation="log2CPMRatio") dat1 <- smoothRatiosByChromosome(dat, N=100, chr = c("chr18", "chr19")) dat2 <- smoothRatiosByChromosome(dat, N=100, chr = c("chr18", "chr19"), chrom.level.background = FALSE)
tileCount extends summarizeOverlaps by finding coverage for each fixed window in the whole genome
tileCount( reads, genome, excludeChrs = c("chrM", "M", "Mt", "MT"), windowSize = 50000, step = 10000, mode = IntersectionNotStrict, dataOverSamples = FALSE, ... )tileCount( reads, genome, excludeChrs = c("chrM", "M", "Mt", "MT"), windowSize = 50000, step = 10000, mode = IntersectionNotStrict, dataOverSamples = FALSE, ... )
reads |
A GRanges,
GRangesList (should be one read per list element),
GAlignments,
GAlignmentsList,
GAlignmentPairs or
BamFileList object that represents the data to be
counted by |
genome |
A BSgenome object from/on which to get/set the sequence and metadata information. |
excludeChrs |
A vector of string: chromosomes/scaffolds of no interest for NAD analysis. see summarizeOverlaps. default is countByOverlaps, alia of countOverlaps(features, reads, ignore.strand=ignore.strand) |
windowSize |
numeric(1) or integer(1). Size of the windows. |
step |
numeric(1) or integer(1). Step of generating silding windows. |
mode |
One of the pre-defined count methods. |
dataOverSamples |
logical(1). Data over several samples when use GRangesList as input. |
... |
Additional arguments passed to
|
A RangedSummarizedExperiment object. The assays slot holds the counts, rowRanges holds the annotation from the sliding widows of genome. metadata contains lib.size.chrom for holding chromosome-level sequence depth
Jianhong Ou, Haibo Liu, Herve Pages and Julie Zhu
if (interactive()) { fls <- list.files(system.file("extdata", package="NADfinder"), recursive=FALSE, pattern="*bam$", full=TRUE) names(fls) <- basename(fls) if (!require(BSgenome.Mmusculus.UCSC.mm10)) { if (!requireNamespace("BiocManager", quietly=TRUE)) install.packages("BiocManager") BiocManager::install("BSgenome.Mmusculus.UCSC.mm10") library(BSgenome.Mmusculus.UCSC.mm10) } se <- tileCount(reads = fls, genome = Mmusculus, excludeChrs = c("chrM", paste0("chr", c(1:17,19)), "chrX", "chrY"), windowSize=50000, step=10000) }if (interactive()) { fls <- list.files(system.file("extdata", package="NADfinder"), recursive=FALSE, pattern="*bam$", full=TRUE) names(fls) <- basename(fls) if (!require(BSgenome.Mmusculus.UCSC.mm10)) { if (!requireNamespace("BiocManager", quietly=TRUE)) install.packages("BiocManager") BiocManager::install("BSgenome.Mmusculus.UCSC.mm10") library(BSgenome.Mmusculus.UCSC.mm10) } se <- tileCount(reads = fls, genome = Mmusculus, excludeChrs = c("chrM", paste0("chr", c(1:17,19)), "chrX", "chrY"), windowSize=50000, step=10000) }
Perform overlap queries between reads and genome by sliding windows Count reads over sliding windows.
if (interactive()) fls <- list.files(system.file("extdata", package="NADfinder"), recursive=FALSE, pattern="*bam$", full=TRUE) names(fls) <- basename(fls)
se <- tileCount2(reads = fls, windowSize=50000, step=10000)
tileCount2( reads, fragment.length = 100, windowSize = 50000, restrict = paste0("chr", c(1:19, "X", "Y")), step = 1000, filter = 0, pe = "both" ) tileCount2( reads, fragment.length = 100, windowSize = 50000, restrict = paste0("chr", c(1:19, "X", "Y")), step = 1000, filter = 0, pe = "both" )tileCount2( reads, fragment.length = 100, windowSize = 50000, restrict = paste0("chr", c(1:19, "X", "Y")), step = 1000, filter = 0, pe = "both" ) tileCount2( reads, fragment.length = 100, windowSize = 50000, restrict = paste0("chr", c(1:19, "X", "Y")), step = 1000, filter = 0, pe = "both" )
reads |
An object that represents the names and path of the bam files to be counted. If reads are more than 1 bam files, it should be a vector of character with full path. This function now works for paired end reads |
fragment.length |
integer(1). An integer scalar or a list of two integer scalars/vectors, containing the average length(s) of the sequenced fragments in each libary. |
windowSize |
numeric(1) or integer(1). Size of the windows. |
restrict |
restrict to a set of chromosomes, default to mouse chromosomes. |
step |
numeric(1) or integer(1). Step of generating silding windows. |
filter |
default to 0 without filtering. An integer scalar for the minimum count sum across libraries for each window |
pe |
a character string indicating whether paired-end data is present; set to "none", "both", "first" or "second" |
A RangedSummarizedExperiment object with chromosome-level depth The assays slot holds the counts, rowRanges holds the annotation from the sliding widows of genome. metadata contains lib.size.chrom for holding chromosome-level sequence depth
Jun Yu,Hervé Pagès and Julie Zhu
if (interactive()) { fls <- list.files(system.file("extdata", package="NADfinder"), recursive=FALSE, pattern="*bam$", full=TRUE) names(fls) <- basename(fls) se <- tileCount2(reads = fls, windowSize=50000, step=10000) }if (interactive()) { fls <- list.files(system.file("extdata", package="NADfinder"), recursive=FALSE, pattern="*bam$", full=TRUE) names(fls) <- basename(fls) se <- tileCount2(reads = fls, windowSize=50000, step=10000) }
calculate the log2 ratios, log2 cpm (count per million) ratios, or log2 odds ratios for nucleolus vs genome. pseudo-count will be used to avoid x/0 or log(0).
transformData( A, B, seqnames.A, seqnames.B, pseudo.count = 1L, transformation = c("log2OddsRatio", "log2CPMRatio", "log2Ratio"), chrom.level.lib = TRUE, lib.size.A, lib.size.B )transformData( A, B, seqnames.A, seqnames.B, pseudo.count = 1L, transformation = c("log2OddsRatio", "log2CPMRatio", "log2Ratio"), chrom.level.lib = TRUE, lib.size.A, lib.size.B )
A, B
|
window-level counts for nucleolus and genome, extracted from the assays of the output of the tileCounts function |
seqnames.A, seqnames.B
|
seqnames, extracted from the rowRanges of the ouput of the tileCounts function |
pseudo.count |
pseudo-count will be used to aviod x/0 or log0, defult to 1. |
transformation |
transformation type |
chrom.level.lib |
indicating whether calculating CPM or odds using sequence depth of the whole genome or the corresponding chromosome |
lib.size.A, lib.size.B
|
library size for A and B. these two dataframes contain chromosome-level sequence depth for the chromosomes, which can be extracted from the metadata of the output of the tileCounts function |
a numeric vector of log2 ratios, log2 CPM ratios or log2 odds ratios.
Julie Zhu
transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2OddsRatio", chrom.level.lib = FALSE, lib.size.A = cbind(c("chr1", "chr2"), c(10000, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 12000))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2CPMRatio", chrom.level.lib = FALSE, lib.size.A = cbind(c("chr1", "chr2"), c(10000, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 12000))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2CPMRatio", chrom.level.lib = TRUE, lib.size.A = cbind(c("chr1", "chr2"), c(100, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 200))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2OddsRatio", chrom.level.lib = TRUE, lib.size.A = cbind(c("chr1", "chr2"), c(100, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 200))) transformData(seq_len(10), 10:1, transformation = "log2Ratio")transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2OddsRatio", chrom.level.lib = FALSE, lib.size.A = cbind(c("chr1", "chr2"), c(10000, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 12000))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2CPMRatio", chrom.level.lib = FALSE, lib.size.A = cbind(c("chr1", "chr2"), c(10000, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 12000))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2CPMRatio", chrom.level.lib = TRUE, lib.size.A = cbind(c("chr1", "chr2"), c(100, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 200))) transformData(seq_len(10), 10:1, seqnames.A = Rle(c("chr1", "chr2" ) , c(5,5)), Rle(c("chr1", "chr2" ) , c(5,5)), transformation = "log2OddsRatio", chrom.level.lib = TRUE, lib.size.A = cbind(c("chr1", "chr2"), c(100, 12000)), lib.size.B = cbind(c("chr1", "chr2"), c(10000, 200))) transformData(seq_len(10), 10:1, transformation = "log2Ratio")
Filter the peaks by pvalue and trim the range of peaks for an NAD or ChIP-seq experiment without biological replicates.
trimPeaks( se, cutoffAdjPvalue = 0.05, padjust.method = "BH", backgroundPercentage = 0.25, countFilter = 1000, ratioAssay = "ratio", backgroundCorrectedAssay = "bcRatio", smoothedRatioAssay = "smoothedRatio", zscoreAssay = "zscore" )trimPeaks( se, cutoffAdjPvalue = 0.05, padjust.method = "BH", backgroundPercentage = 0.25, countFilter = 1000, ratioAssay = "ratio", backgroundCorrectedAssay = "bcRatio", smoothedRatioAssay = "smoothedRatio", zscoreAssay = "zscore" )
se |
An object of RangedSummarizedExperiment with assays of raw counts, ratios, background corrected ratios, smoothed ratios and z-scores. It should be an element of the output of smoothRatiosByChromosome |
cutoffAdjPvalue |
numeric(1). Cutoff of adjusted p-value. |
padjust.method |
character(1). The method to use for adjusting p-values, which is passed to p.adjust function |
backgroundPercentage |
numeric(1). Cutoff value for the peaks height. |
countFilter |
numeric(1) or integer(1). Cutoff value for mean of raw reads count of the Nucleolar/ChIP samples in each window. |
ratioAssay |
character(1). The name of assay in se, which store the values to be smoothed. |
backgroundCorrectedAssay, smoothedRatioAssay, zscoreAssay
|
Assays names for background-corrected ratios, smoothed ratios and z-scores based on background corrected ratios. |
An object of GRanges.
data(single.count) se <- single.count dat <- log2se(se, nucleolusCols="N18.subsampled.srt.bam", genomeCols="G18.subsampled.srt.bam", transformation="log2CPMRatio") ## Smooth the ratios for each chromosome. dat <- smoothRatiosByChromosome(dat, N=100, chr=c("chr18","chr19")) peaks <- trimPeaks(dat[["chr18"]], backgroundPercentage=.25, cutoffAdjPvalue=0.05, countFilter=1000)data(single.count) se <- single.count dat <- log2se(se, nucleolusCols="N18.subsampled.srt.bam", genomeCols="G18.subsampled.srt.bam", transformation="log2CPMRatio") ## Smooth the ratios for each chromosome. dat <- smoothRatiosByChromosome(dat, N=100, chr=c("chr18","chr19")) peaks <- trimPeaks(dat[["chr18"]], backgroundPercentage=.25, cutoffAdjPvalue=0.05, countFilter=1000)
Counts data for chromosome 18 for an expriment with triplicates
Calculate the z-scores over the background distribution.
zscoreOverBck(ratios, backgroundPercentage = 0.25)zscoreOverBck(ratios, backgroundPercentage = 0.25)
ratios |
A numeric vector containing the transformed, background corrected and smoothed ratios in each window. |
backgroundPercentage |
numeric(1). Low percentile for background distribution. |
A vector of numeric. Z-scores.
Jianhong Ou and Julie Zhu
r <- runif(200) zscoreOverBck(r)r <- runif(200) zscoreOverBck(r)