This tool takes an alignment of reads or fragments as input (BAM file) and generates a coverage track (bigWig or bedGraph) as output. The coverage is calculated as the number of reads per bin, where bins are short consecutive counting windows of a defined size. It is possible to extended the length of the reads to better reflect the actual fragment length. bamCoverage offers normalization by scaling factor, Reads Per Kilobase per Million mapped reads (RPKM), and 1x depth (reads per genome coverage, RPGC).

usage: An example usage is: bamCoverage -b signal.bam -o
Required arguments
--bam, -b BAM file to process
--outFileName, -o
 Output file name.
--outFileFormat=bigwig, -of=bigwig

Output file type. Either “bigwig” or “bedgraph”.

Possible choices: bigwig, bedgraph

Optional arguments
 Indicate a number that you would like to use. When used in combination with –normalizeTo1x or –normalizeUsingRPKM, the computed scaling factor will be multiplied by the given scale factor.
--MNase=False Determine nucleosome positions from MNase-seq data. Only 3 nucleotides at the center of each fragment are counted. The fragment ends are defined by the two mate reads. Only fragment lengthsbetween 130 - 200 bp are considered to avoid dinucleosomes or other artifacts.*NOTE*: Requires paired-end data. A bin size of 1 is recommended.
--version show program’s version number and exit
--binSize=50, -bs=50
 Size of the bins, in bases, for the output of the bigwig/bedgraph file.
--region, -r Region of the genome to limit the operation to - this is useful when testing parameters to reduce the computing time. The format is chr:start:end, for example –region chr10 or –region chr10:456700:891000.
--numberOfProcessors=max/2, -p=max/2
 Number of processors to use. Type “max/2” to use half the maximum number of processors or “max” to use all available processors.
--verbose=False, -v=False
 Set to see processing messages.
Read coverage normalization options
 Report read coverage normalized to 1x sequencing depth (also known as Reads Per Genomic Content (RPGC)). Sequencing depth is defined as: (total number of mapped reads * fragment length) / effective genome size. The scaling factor used is the inverse of the sequencing depth computed for the sample to match the 1x coverage. To use this option, the effective genome size has to be indicated after the option. The effective genome size is the portion of the genome that is mappable. Large fractions of the genome are stretches of NNNN that should be discarded. Also, if repetitive regions were not included in the mapping of reads, the effective genome size needs to be adjusted accordingly. Common values are: mm9: 2,150,570,000; hg19:2,451,960,000; dm3:121,400,000 and ce10:93,260,000. See Table 2 of or for several effective genome sizes.
 Use Reads Per Kilobase per Million reads to normalize the number of reads per bin. The formula is: RPKM (per bin) = number of reads per bin / ( number of mapped reads (in millions) * bin length (kb) ). Each read is considered independently,if you want to only count either of the mate pairs inpaired-end data, use the –samFlag option.
--ignoreForNormalization, -ignore
 A list of space-delimited chromosome names containing those chromosomes that should be excluded for computing the normalization. This is useful when considering samples with unequal coverage across chromosomes, like male samples. An usage examples is –ignoreForNormalization chrX chrM.
--skipNonCoveredRegions=False, --skipNAs=False
 This parameter determines if non-covered regions (regions without overlapping reads) in a BAM file should be skipped. The default is to treat those regions as having a value of zero. The decision to skip non-covered regions depends on the interpretation of the data. Non-covered regions may represent, for example, repetitive regions that should be skipped.
--smoothLength The smooth length defines a window, larger than the binSize, to average the number of reads. For example, if the –binSize is set to 20 and the –smoothLength is set to 60, then, for each bin, the average of the bin and its left and right neighbors is considered. Any value smaller than –binSize will be ignored and no smoothing will be applied.
Read processing options
--extendReads=False, -e=False
 This parameter allows the extension of reads to fragment size. If set, each read is extended, without exception. *NOTE*: This feature is generally NOT recommended for spliced-read data, such as RNA-seq, as it would extend reads over skipped regions. *Single-end*: Requires a user specified value for the final fragment length. Reads that already exceed this fragment length will not be extended. *Paired-end*: Reads with mates are always extended to match the fragment size defined by the two read mates. Unmated reads, mate reads that map too far apart (>4x fragment length) or even map to different chromosomes are treated like singe-end reads. The input of a fragment length value is optional. If no value is specified, it is estimated from the data (mean of the fragment size of all mate reads).
 If set, reads that have the same orientation and start position will be considered only once. If reads are paired, the mate’s position also has to coincide to ignore a read.
 If set, only reads that have a mapping quality score of at least this are considered.
 By adding this option, reads are centered with respect to the fragment length. For paired-end data, the read is centered at the fragment length defined by the two ends of the fragment. For single-end data, the given fragment length is used. This option is useful to get a sharper signal around enriched regions.
 Include reads based on the SAM flag. For example, to get only reads that are the first mate, use a flag of 64. This is useful to count properly paired reads only once, as otherwise the second mate will be also considered for the coverage.
 Exclude reads based on the SAM flag. For example, to get only reads that map to the forward strand, use –samFlagExclude 16, where 16 is the SAM flag for reads that map to the reverse strand.

Usage hints

  • A smaller bin size value will result in a higher resolution of the coverage track but also in a larger file size.
  • The 1x normalization (RPGC) requires the input of a value for the effective genome size, which is the mappable part of the reference genome. Of course, this value is species-specific. The command line help of this tool offers suggestions for a number of model species.
  • It might be useful for some studies to exclude certain chromosomes in order to avoid biases, e.g. chromosome X, as male mice contain a pair of each autosome, but usually only a single X chromosome.
  • By default, the read length is NOT extended! This is the preferred setting for spliced-read data like RNA-seq, where one usually wants to rely on the detected read locations only. A read extension would neglect potential splice sites in the unmapped part of the fragment. Other data, e.g. Chip-seq, where fragments are known to map contiguously, should be processed with read extension (--extendReads [INT]).
  • For paired-end data, the fragment length is generally defined by the two read mates. The user provided fragment length is only used as a fallback for singletons or mate reads that map too far apart (with a distance greater than four times the fragment length or are located on different chromosomes).


If you already normalized for GC bias using correctGCbias, you should absolutely NOT set the parameter --ignoreDuplicates!

Usage example

This is an example using additional options (smaller bin size for higher resolution, normalizing coverage to 1x mouse genome size, excluding chromosome X during the normalization step, and extending reads):

$ bamCoverage --bam reads.bam -o
   --binSize 10
   --normalizeTo1x 2150570000
   --ignoreForNormalization chrX


bamCoverage is also available in deepTools Galaxy: