Converts paired-end BAM to BED12 format, based on bedtools
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README.rst

pairedBamToBed12

https://travis-ci.org/Population-Transcriptomics/pairedBamToBed12.svg?branch=pairedbamtobed12

pairedBamToBed12 converts properly paired BAM alignments to BED12 format. Typical proper pairs will be represented by a 2 blocks BED12 entry. Additional blocks are produced when an alignment contains long deletion (CIGAR N-op). Thickness indicates the first read of the pair. The BAM input file must be grouped/sorted by query name (not alignment position).

Read 1:   >>>>>>>>>>>>
Read 2:                     <<<<<<<<<<<<<-----<<<<<<<
The pair: >>>>>>>>>>>>------>>>>>>>>>>>>>----->>>>>>>

Installation

In brief: download the source code, unpack it and enter the main source directory, type make and a pairedBamToBed12 executable file will appear the bin directory. Test it with make test. See below for an example.

wget https://github.com/Population-Transcriptomics/pairedBamToBed12/archive/pairedbamtobed12.zip
unzip pairedbamtobed12.zip
cd pairedBamToBed12-pairedbamtobed12
make
make test
ls bin/pairedBamToBed12

Usage and option summary

Usage:

pairedBamToBed12 [OPTIONS] -i <BAM>
.. tabularcolumns:: |p{4.5cm}|p{8.5cm}|

Option Description
-dblock Triggers the creation of a new block when an alignment contains short deletion from reference (CIGAR D-op).
-color An R,G,B string for the color used with BED12 format. Default is (255,0,0).
-extraG Ignore G mismatches on first bases (experimental option for use on CAGE alignments with BWA aln).
-nsep A string after which the read names are allowed to differ. Default is ___. Give an improbable value like 'nothankyou' to turn off.
-qual The minimum (inclusive) mapQ sum for reporting the paired BAM into a BED12. Default is 0.
-x Optional filename where unprocessed mapped pairs can be stored.

Default behavior

By default it processes a properly paired pair of reads into a single BED12 line, where the start and end positions are the 5′ end of Read 1 and the 3′ end of Read 2. The BED12 blocks are used to indicate positions where the reads match, and the thick part indicates where is the contribution of Read 1. The relative orientation of the mate pairs must be forward/reverse (which is the standard in most libraries prepared for the Illumina platform).

Note

The BAM file must be sorted by read name.

Note

Reads that are not followed by their mate or not properly paired will be skipped.

$ pairedBamToBed12 -i 1proper-pair.bam
chr1  50053297        50053480        M00528:19:000000000-A88YD:1:1101:2241:12366     0       +       50053297        50053324        255,0,0 2       27,21   0,162

Usage with transcriptome libraries

In transcriptome analysis, the BED12 entries produced by pairedBamToBed12 represent the minimal information about a cDNA that was given by a read pair.

pairedBamToBed12 was created for the analysis of CAGEscan libraries, which are paired-end directional libraries of random-primed 5′ cDNAs. The BED12 files are used to assemble CAGEscan clusters that combine all the pairs where the 5′ end is in the same transcript start site peak, thus providing approximate rudimentary transcript models for each peak. A typical analysis can be found in Kratz et al., 2014.

This BED12 format is also supported in RIKEN's Zenbu genome browser, where one can load data in this format and visualise it either as genome intervals or as expression histograms.

Note

BWA has a bug that will set the properly paired flag for reads where one mate is aligned very near the end of a chromosome and the other is aligned very near the beginning of the next chromosome, when the -a option of sampe is large. However, for CAGEscan, large numbers are necessary to span whole gene loci. It is therefore recommended to sanitise the output of BWA with SAMtools, using its fixmate command, that corrects the properly paired flag since version 1.0.

Note

CAGE methods sometimes add an extra G at the beginning of the cDNAs (see http://population-transcriptomics.org/nanoCAGE/#extra-G). This leads to 1-base shifts of some TSS peaks. From version 1.2, pairedBamToBed12 provides an experimental option, -extraG to shift the start or end (according to the strand) of the output of one base when a G mismatch is detected on the first base of Read1. A more detailed description of the problem may be found in the supplemental material of the FANTOM3 paper. The implementation here is very naive and incomplete, and was tested only on data produced by BWA's sampe command. It relies on the MD flag and does not understand clipping. Thus, the -extraG option available here is not entierly satisfactory and may be removed in the future. A better approach for instance would be to post-process the BAM file instead of implementing a correction here.

Advantages and limitations in comparison with bedtools bamtobed

The advantage compared to bedtools bamtobed -split is that pairedBamToBed12 reports the whole pair on a single line, and the advantage compared with bedtools bamtobed -bedpe, is that it reports spliced alignments.

The limitation of pairedbamtobed12 is that it only pertains to pairs mapped on the same chromosome and is therefore unfit for representing gene fusions or interchromosomal interactions.

Copyright, authorship and license

pairedBamToBed12 is distrubuted under the GNU General Public License version 2.

The tool pairedBamToBed12 is copyright 2013~2015 RIKEN. It was originally written by Nicolas Bertin as an addition to Bedtools 2.11.1. It was then ported to Bedtools 2.21.0 by Mickaël Mendez, and then finally forked from the Bedtools source as a stand-alone program by Charles Plessy. The documentation was written by NB, MM and CP, and the regression tests were implemented by MM and CP.

Bedtools is copyright Aaron Quinlan and others.