lumpy sv
v0.3.1
NOTE: This is LUMPY 0.2.13 with additional changes to allow lumpyexpress to function when the main file is a CRAM, not a BAM. The splitters and discordants must still be BAM files as LUMPY itself doesn't yet support CRAM as an input. This requires the hexdump command be available.
For questions and discussion about LUMPY please visit the forum at:
https://groups.google.com/forum/#!forum/lumpy-discuss
A probabilistic framework for structural variant discovery.
Ryan M Layer, Colby Chiang, Aaron R Quinlan, and Ira M Hall. 2014. "LUMPY: a Probabilistic Framework for Structural Variant Discovery." Genome Biology 15 (6): R84. doi:10.1186/gb-2014-15-6-r84.
Note that smoove is the recommended way to run lumpy as it collects the
best-practices of lumpy and associated tools and will have a shorter run-time and lower false-positive rate than
lumpyexpress described below.
Download and install
git clone --recursive https://github.com/arq5x/lumpy-sv.git
cd lumpy-sv
make
cp bin/* /usr/local/bin/.
Run LUMPY Express
lumpyexpress
-B my.bam
-S my.splitters.bam
-D my.discordants.bam
-o output.vcf
Default method to install:
git clone --recursive [email protected]:arq5x/lumpy-sv.git
cd lumpy-sv
make
cp bin/* /usr/local/bin/.
Installing with costom zlib (gzopen64 compile error):
git clone --recursive [email protected]:arq5x/lumpy-sv.git
cd lumpy-sv
export ZLIB_PATH="/usr/lib/x86_64-linux-gnu/"; #when /usr/lib/x86_64-linux-gnu/libz.so
make
cp bin/* /usr/local/bin/.
Automated breakpoint detection for standard analyses.
usage: lumpyexpress [options]
Required arguments
-B FILE coordinate-sorted BAM file(s) (comma separated)
-S FILE split reads BAM file(s) (comma separated)
-D FILE discordant reads BAM files(s) (comma separated)
Optional arguments
-o STR output [fullBam.bam.vcf]
-x FILE BED file to exclude
-P output probability curves for each variant
-m INT minimum sample weight for a call [4]
-r FLOAT trim threshold [0]
-T DIR temp directory [./output_prefix.XXXXXXXXXXXX]
-k keep temporary files
-K FILE path to lumpyexpress.config file
(default: same directory as lumpyexpress)
-v verbose
-h show this message
LUMPY Express runs several external program whose paths are specified in scripts/lumpyexpress.config. This config must reside in the same directory as lumpyexpress, or be specified explicitly with the -K flag.
The installation Makefile auto-generates a lumpyexpress.config file and places it in the "bin" directory.
LUMPY Express expects BWA-MEM aligned BAM files as input. It automatically parses sample, library, and read group information using the @RG tags in the BAM header. Each BAM file is expected to contain exactly one sample.
The minimum input is a coordinate-sorted BAM file (-B), from which LUMPY Express extracts splitters and discordants using SAMBLASTER before running LUMPY. Optionally, users may supply coordinate-sorted splitter (-S) and discordant (-D) BAM files which will bypass SAMBLASTER extraction for faster analysis.
LUMPY Express produces a VCF file according to VCF spec 4.2.
Flexible and customizable breakpoint detection for advanced users.
usage: lumpy [options]
Options
-g Genome file (defines chromosome order)
-e Show evidence for each call
-w File read windows size (default 1000000)
-mw minimum weight across all samples for a call
-msw minimum per-sample weight for a call
-tt trim threshold
-x exclude file bed file
-t temp file prefix, must be to a writeable directory
-P output probability curve for each variant
-b output as BEDPE instead of VCF
-sr bam_file:,
id:,
back_distance:,
min_mapping_threshold:,
weight:,
min_clip:,
read_group:
-pe bam_file:,
id:,
histo_file:,
mean:,
stdev:,
read_length:,
min_non_overlap:,
discordant_z:,
back_distance:,
min_mapping_threshold:,
weight:,
read_group:
-bedpe bedpe_file:,
id:,
weight:
We recommend aligning data with SpeedSeq, which performs BWA-MEM alignment, marks duplicates and extracts split and discordant read-pairs.
speedseq align -R "@RGtID:idtSM:sampletLB:lib"
human_g1k_v37.fasta
sample.1.fq
sample.2.fq
Otherwise, data may be aligned with BWA-MEM.
# Align the data
bwa mem -R "@RGtID:idtSM:sampletLB:lib" human_g1k_v37.fasta sample.1.fq sample.2.fq
| samblaster --excludeDups --addMateTags --maxSplitCount 2 --minNonOverlap 20
| samtools view -S -b -
> sample.bam
# Extract the discordant paired-end alignments.
samtools view -b -F 1294 sample.bam > sample.discordants.unsorted.bam
# Extract the split-read alignments
samtools view -h sample.bam
| scripts/extractSplitReads_BwaMem -i stdin
| samtools view -Sb -
> sample.splitters.unsorted.bam
# Sort both alignments
samtools sort sample.discordants.unsorted.bam sample.discordants
samtools sort sample.splitters.unsorted.bam sample.splitters
LUMPY has two distinct execution alternatives. LUMPY Express is a simplified wrapper for standard analyses. LUMPY (traditional) is more customizable, for advanced users and specialized experiments.
Run LUMPY Express on a single sample with pre-extracted splitters and discordants
lumpyexpress
-B sample.bam
-S sample.splitters.bam
-D sample.discordants.bam
-o sample.vcf
Run LUMPY Express jointly on multiple samples with pre-extracted splitters and discordants
lumpyexpress
-B sample1.bam,sample2.bam,sample3.bam
-S sample1.splitters.bam,sample2.splitters.bam,sample3.splitters.bam
-D sample1.discordants.bam,sample2.discordants.bam,sample3.discordants.bam
-o multi_sample.vcf
Run LUMPY Express on a tumor-normal pair
lumpyexpress
-B tumor.bam,normal.bam
-S tumor.splitters.bam,normal.splitters.bam
-D tumor.discordants.bam,normal.discordants.bam
-o tumor_normal.vcf
First, generate empirical insert size statistics on each library in the BAM file
samtools view -r readgroup1 sample.bam
| tail -n+100000
| scripts/pairend_distro.py
-r 101
-X 4
-N 10000
-o sample.lib1.histo
The above script (scripts/pairend_distro.py) will display mean and stdev to screen. For these examples we will assume the mean is 500 and the stdev is 50.
Run LUMPY with paired-end and split-reads.
lumpy
-mw 4
-tt 0
-pe id:sample,bam_file:sample.discordants.bam,histo_file:sample.lib1.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-sr id:sample,bam_file:sample.splitters.bam,back_distance:10,weight:1,min_mapping_threshold:20
> sample.vcf
Run LUMPY on a BAM file with multiple libraries.
lumpy
-mw 4
-tt 0
-pe id:sample,read_group:rg1,bam_file:sample.discordants.bam,histo_file:sample.lib1.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-pe id:sample,read_group:rg2,bam_file:sample.discordants.bam,histo_file:sample.lib2.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-sr id:sample,bam_file:sample.splitters.bam,back_distance:10,weight:1,min_mapping_threshold:20
> sample.vcf
Run LUMPY on multiple samples with multiple libraries.
lumpy
-mw 4
-tt 0
-pe id:sample1,bam_file:sample1.discordants.bam,read_group:rg1,read_group:rg2,histo_file:sample1.lib1.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-pe id:sample1,bam_file:sample1.discordants.bam,read_group:rg3,histo_file:sample1.lib2.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-pe id:sample2,bam_file:sample2.discordants.bam,read_group:rg4,histo_file:sample2.lib1.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,min_mapping_threshold:20
-sr id:sample1,bam_file:sample1.splitters.bam,back_distance:10,weight:1,min_mapping_threshold:20
-sr id:sample2,bam_file:sample2.splitters.bam,back_distance:10,weight:1,min_mapping_threshold:20
> multi_sample.vcf
Run LUMPY with low complexity regions excluded.
Heng Li provides a set of low complexity regions in the supplementary information of his paper, "Toward better
understanding of artifacts in variant calling from high-coverage samples" at
https://doi.org/10.1093/bioinformatics/btu356.
unzip btu356_Supplementary_Data.zip
unzip btu356-suppl_data.zip
lumpy
-mw 4
-tt 0.0
-x btu356_LCR-hs37d5.bed/btu356_LCR-hs37d5.bed
-pe bam_file:sample.discordants.bam,histo_file:sample.pe.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,id:sample,min_mapping_threshold:1
-sr bam_file:sample.sr.sort.bam,back_distance:10,weight:1,id:sample,min_mapping_threshold:1
> sample.exclude.vcf
Run LUMPY with regions of very high coverage excluded.
We can direct lumpy to ignore certain regions by using the
exclude region option. In this example we find and then
exclude regions that have very high coverage. First we
use the get_coverages.py script to find the min, max, and
mean coverages of the the sr and pe bam files, and to
create coverage profiles for both files.
python ../scripts/get_coverages.py
sample.pe.sort.bam
sample.sr.sort.bam
# sample.pe.sort.bam.coverage min:1 max:14 mean(non-zero):2.35557521272
# sample.sr.sort.bam.coverage min:1 max:7 mean(non-zero):1.08945936729
From this output, we will choose to exclude regions that have more than 10x coverage. To create the exclude file we will use the get_exclude_regions.py script to create the exclude.bed file
python ../scripts/get_exclude_regions.py
10
exclude.bed
sample.pe.sort.bam
sample.sr.sort.bam
We now rerun lumpy with the exclude (-x) option
lumpy
-mw 4
-tt 0.0
-x exclude.bed
-pe bam_file:sample.discordants.bam,histo_file:sample.pe.histo,mean:500,stdev:50,read_length:101,min_non_overlap:101,discordant_z:5,back_distance:10,weight:1,id:sample,min_mapping_threshold:1
-sr bam_file:sample.sr.sort.bam,back_distance:10,weight:1,id:sample,min_mapping_threshold:1
> sample.exclude.vcf
SVTyper can call genotypes on LUMPY output VCF files using a Bayesian maximum likelihood algorithm.
svtyper
-B sample.bam
-S sample.splitters.bam
-i sample.vcf
> sample.gt.vcf
The test/test.sh script executes lumpy against several simulated data sets
and compares the results to the known correct result. The sample data sets can
be found at http://layerlab.org/lumpy/data.tar.gz. This tar ball should be
extracted into the top-level lumpy directory. The script test/test.sh checks
for the the existence of this directory before running LUMPY.
All of the bam files that lumpy processes must be position sorted. To check if your bams are sorted correctly, use the check_sorting.py script
python ../scripts/check_sorting.py
pe.pos_sorted.bam
sr.pos_sorted.bam
pe.name_sorted.bam
# pe.pos_sorted.bam
# in order
# sr.pos_sorted.bam
# in order
# pe.name_sorted.bam
# out of order: chr10 102292476 occurred after chr10 102292893
