damid.seq: DamID-Seq pipeline

In damidseq/RDamIDSeq: DamID-seq pipeline (Adapter removal, Mapping to genome, analyzing)

Description

Full DamID-Seq pipeline; includes adapter removal, mapping and enrichment finding.

Usage

damid.seq(raw.file.name, input.format="fastq", multi.core=F,
          exp.name="damid-gatc-sites",
          bin.len=100000, adapt.seq="CGCGGCCGAG", errors=1, qc=T,
          species="BSgenome.Celegans.UCSC.ce10", chr.names = NULL,
          restr.seq="GATC", normali=T, log10t=T, m.hits=1)

Arguments

raw.file.name	A tab-separated file with 2 or 3 columns. See raw-file.txt as example! first column: the path to the raw fastq (fastq.gz) or fasta file (if mapping = FALSE, the file must be a grange .RData object). Second column: name of the sample. Third column: single character ‘s’ for sample and ‘c’ for control (if not set the pipeline does not perform a log2 fold change output). The number of samples ("s") must be the same as the number of controls ("c").
input.format	if "fastq" or "fasta", the pipeline starts with the raw fastq/a(.gz) or fastq/a files after sequencing. If "g", the pipeline starts with the GRange object made by the full pipeline to skip the adapter removal and rbowtie mapping. If "b" the pipeline starts with mapped .bam files
multi.core	if TRUE, bowtie is performed with multiple cores
exp.name	a generic name for your experiment
bin.len	the bin length for the analysis
errors	errors in adapter sequence allowed during adapter removal
adapt.seq	the adapter sequence to be removed before mapping
qc	if TRUE, a raw reads quality control is performed
species	an object of class "BSgenome", the name of the BSgenome object for rbowtie mapping and GATC sites extraction
chr.names	a string vector of chromosome names convention of BSgenome-utils or NULL to take all chromosomes from species
restr.seq	the sequence of the restriction site
normali	if TRUE, the reads are normalized by dividing the reads in a bin by the total reads numbers of the sample
log10t	if TRUE, reads per bin are log transformed in qc plots
m.hits	Number of mapping positions allowed per read for bowtie mapping (1 = only unique mappable reads are mapped)
...	not used.

Details

The tab-separated values in raw.file.name must be provided as shown in the example below to analyze a sample using controls. Column names can be chosen freely.

FilePath/Name	sampleName	groupIndex
rawfile/sample.fastq.gz	test.sample	"s"
rawfile/contro.fastq.gz	test.control	"c"

The group index can be omited if no sample-control comparison is wished. (Log2 fold change between sample and control will not be plotted)

FilePath/Name	sampleName
rawfile/sample.fastq.gz	test.sample
rawfile/contro.fastq.gz	test.contrl

several files are saved during the pipeline in several folders.

folder name	saved files
cutadapter	cut reads (.fastq/a)
cutadapter	cut read length (.txt)
cutadapter	cut read information (.txt)
bowtie	mapped reads (.bam)
bowtie	bam information (.bam.bai)
bowtie	bam information (.txt)
granges	reads per input (sample) file (.Rdata)
granges	reads per GATC site (.Rdata)
granges	reads per GATC site plus strand (.Rdata)
granges	reads per GATC site minus strand (.Rdata)
granges	reads per GATC fragment (.Rdata)
granges	GATC sites in genome
qc-reports	fastqc report if qc = T (.txt)
qc-reports	correlation between samples: GATC and bin (.txt)
qc-reports	number of reads lost in each step (.txt)
results	correlation and read distribution: GATC and bin (.pdf)
results	read plots: GATC and bin (.pdf)
results	log2 fold change sample / control (.pdf)

Value

A list with 5 GRanges-class objects.

1	$GATC	sequencing reads per GATC site
2	$GATC.plus	sequencing reads per GATC site on the plus strand
3	$GATC.minus	sequencing reads per GATC site on the minus strand
4	$GATC.frag	sequencing reads per GATC fragment
5	$bins	sequencing reads per chosen bin

Author(s)

Dominic Ritler

Examples

damid.seq(raw.file.name = "raw-file.txt", input.format = "fastq",
          multi.core = T, exp.name = "test", bin.len = 100000,
          adapt.seq="CGCGGCCGAG", errors = 1,  qc = F,
          species = "BSgenome.Celegans.UCSC.ce10",
          chr.names = NULL, restr.seq = "GATC", normali = T, log10t = F,
          m.hits = 1)

damidseq/RDamIDSeq documentation built on May 14, 2019, 3:33 p.m.