PostprocessGermlineCNVCalls

Postprocesses the output of GermlineCNVCaller and generates VCFs and denoised copy ratios

Category Copy Number Variant Discovery

Overview

Postprocesses the output of {@link GermlineCNVCaller} and generates VCF files as well as a concatenated denoised copy ratio file.

This tool generates "intervals" and "segments" VCF files that serve complementary purposes. The intervals VCF file provides a detailed listing of the most likely copy-number call for each genomic interval included in the call-set, along with call quality, call genotype, and the phred-scaled posterior probability vector for all integer copy-number states. Given that CNV events often span several consecutive intervals, it may be desirable to coalesce contiguous intervals with the same copy-number call into a constant copy-number segments. This tool further performs segmentation and genotyping by calling a dedicated python script in `gcnvkernel`. The segmentation algorithm further provides various quality metrics for the segment.

For both VCF outputs, the CNV genotype is determined as follows: the alternative allele for a CNV call is either <DEL> or <DUP>, depending on whether the most likely copy-number call is below or above the reference copy-number for the contig. The user may specify the reference copy-number state on autosomal contigs using the argument autosomal-ref-copy-number. The list of allosomal contigs may also be specified via the argument allosomal-contig. All undeclared contigs are assumed to be autosomal. The reference copy-number on an allosomal contig is determined by the sex karyotype of the sample and is set to the pre-determined contig ploidy state fetched from the output calls of {@link DetermineGermlineContigPloidy}.

Finally, the tool concatenates posterior means for denoised copy ratios from all the call shards produced by the {@link GermlineCNVCaller} into a single file.

This tool can also take a VCF specifying breakpoints to be used instead of HMM-derived segmentation using posterior probabilities from the intervals. This functionality enables the calculation of new quality scores using breakpoints derived from another source, as with JointGermlineCNVSegmentation applied to multiple samples. When using this functionality, an input-intervals-vcf from the original PostprocessGermlineCNVCalls call without multi-sample breakpoints should also be provided.

Python environment setup

The computation done by this tool, aside from input data parsing and validation, is performed outside of the Java Virtual Machine and using the gCNV computational python module, namely {@code gcnvkernel}. It is crucial that the user has properly set up a python conda environment with {@code gcnvkernel} and its dependencies installed. If the user intends to run {@link PostprocessGermlineCNVCalls} using one of the official GATK Docker images, the python environment is already set up. Otherwise, the environment must be created and activated as described in the main GATK README.md file.

Advanced users may wish to set the PYTENSOR_FLAGS environment variable to override the GATK PyTensor configuration. For example, by running PYTENSOR_FLAGS="base_compiledir=PATH/TO/BASE_COMPILEDIR" gatk DetermineGermlineContigPloidy ..., users can specify the PyTensor compilation directory (which is set to $HOME/.pytensor by default). See PyTensor documentation at https://pytensor.readthedocs.io/en/latest/library/config.html.

Required inputs:

A list of paths to {@link GermlineCNVCaller} calls shards
A list of paths to {@link GermlineCNVCaller} model shards
Path to the output calls of {@link DetermineGermlineContigPloidy}
Index of the sample in the call-set (which is expected to be the same across all shards)
Output path for writing the intervals VCF
Output path for writing the segments VCF
Output path for writing the concatenated denoised copy ratios

The calls or model shards can be specified in arbitrary order.

Usage examples

   gatk PostprocessGermlineCNVCalls \
     --calls-shard-path path/to/shard_1-calls
     --calls-shard-path path/to/shard_2-calls
     --model-shard-path path/to/shard_1-model
     --model-shard-path path/to/shard_2-model
     --sample-index 0
     --autosomal-ref-copy-number 2
     --allosomal-contig X
     --allosomal-contig Y
     --output-genotyped-intervals sample_0_genotyped_intervals.vcf
     --output-genotyped-segments sample_0_genotyped_segments.vcf
     --output-denoised-copy-ratios sample_0_denoised_copy_ratios.tsv

   gatk PostprocessGermlineCNVCalls \
     --calls-shard-path path/to/shard_1-calls
     --calls-shard-path path/to/shard_2-calls
     --model-shard-path path/to/shard_1-model
     --model-shard-path path/to/shard_2-model
     --sample-index 0
     --autosomal-ref-copy-number 2
     --allosomal-contig X
     --allosomal-contig Y
     --input-intervals-vcf sample_0_genotyped_intervals.vcf
     --clustered-breakpoints cohort_breakpoints.vcf
     --output-genotyped-intervals sample_0_genotyped_intervals.clustered.vcf
     --output-genotyped-segments sample_0_genotyped_segments.clustered.vcf
     --output-denoised-copy-ratios sample_0_denoised_copy_ratios.clustered.tsv
     -R reference.fasta

@author Mehrtash Babadi <mehrtash@broadinstitute.org> @author Andrey Smirnov <asmirnov@broadinstitute.org>

Additional Information

Read filters

This Read Filter is automatically applied to the data by the Engine before processing by PostprocessGermlineCNVCalls.

WellformedReadFilter

PostprocessGermlineCNVCalls specific arguments

This table summarizes the command-line arguments that are specific to this tool. For more details on each argument, see the list further down below the table or click on an argument name to jump directly to that entry in the list.

Argument name(s)	Default value	Summary
Required Arguments
--calls-shard-path		List of paths to GermlineCNVCaller call directories.
--contig-ploidy-calls		Path to contig-ploidy calls directory (output of DetermineGermlineContigPloidy).
--model-shard-path		List of paths to GermlineCNVCaller model directories.
--output-denoised-copy-ratios		Output denoised copy ratio file.
--output-genotyped-intervals		Output intervals VCF file.
--output-genotyped-segments		Output segments VCF file.
Optional Tool Arguments
--allosomal-contig		Contigs to treat as allosomal (i.e. choose their reference copy-number allele according to the sample karyotype).
--arguments_file		read one or more arguments files and add them to the command line
--autosomal-ref-copy-number	2	Reference copy-number on autosomal intervals.
--cloud-index-prefetch-buffer -CIPB	-1	Size of the cloud-only prefetch buffer (in MB; 0 to disable). Defaults to cloudPrefetchBuffer if unset.
--cloud-prefetch-buffer -CPB	40	Size of the cloud-only prefetch buffer (in MB; 0 to disable).
--clustered-breakpoints		VCF with clustered breakpoints and copy number calls for all samples, can be generated with GATK JointGermlineCNVSegmentation tool
--disable-bam-index-caching -DBIC	false	If true, don't cache bam indexes, this will reduce memory requirements but may harm performance if many intervals are specified. Caching is automatically disabled if there are no intervals specified.
--disable-sequence-dictionary-validation	false	If specified, do not check the sequence dictionaries from our inputs for compatibility. Use at your own risk!
--duplication-qs-threshold	50	Filter out duplications with quality lower than this.
--gcs-max-retries -gcs-retries	20	If the GCS bucket channel errors out, how many times it will attempt to re-initiate the connection
--gcs-project-for-requester-pays		Project to bill when accessing "requester pays" buckets. If unset, these buckets cannot be accessed. User must have storage.buckets.get permission on the bucket being accessed.
--help -h	false	display the help message
--het-deletion-qs-threshold	100	Filter out heterozygous deletions with quality lower than this.
--hom-deletion-qs-threshold	400	Filter out homozygous deletions with quality lower than this.
--input-intervals-vcf		Input VCF with combined intervals for all samples
--interval-merging-rule -imr	ALL	Interval merging rule for abutting intervals
--intervals -L		One or more genomic intervals over which to operate
--reference -R		Reference sequence
--sample-index	0	Sample index in the call-set (must be contained in all shards).
--site-frequency-threshold	0.01	Filter out variants with site frequency higher than this.
--sites-only-vcf-output	false	If true, don't emit genotype fields when writing vcf file output.
--version	false	display the version number for this tool
Optional Common Arguments
--add-output-sam-program-record	true	If true, adds a PG tag to created SAM/BAM/CRAM files.
--add-output-vcf-command-line	true	If true, adds a command line header line to created VCF files.
--create-output-bam-index -OBI	true	If true, create a BAM/CRAM index when writing a coordinate-sorted BAM/CRAM file.
--create-output-bam-md5 -OBM	false	If true, create a MD5 digest for any BAM/SAM/CRAM file created
--create-output-variant-index -OVI	true	If true, create a VCF index when writing a coordinate-sorted VCF file.
--create-output-variant-md5 -OVM	false	If true, create a a MD5 digest any VCF file created.
--disable-read-filter -DF		Read filters to be disabled before analysis
--disable-tool-default-read-filters	false	Disable all tool default read filters (WARNING: many tools will not function correctly without their default read filters on)
--exclude-intervals -XL		One or more genomic intervals to exclude from processing
--gatk-config-file		A configuration file to use with the GATK.
--input -I		BAM/SAM/CRAM file containing reads
--interval-exclusion-padding -ixp	0	Amount of padding (in bp) to add to each interval you are excluding.
--interval-padding -ip	0	Amount of padding (in bp) to add to each interval you are including.
--interval-set-rule -isr	UNION	Set merging approach to use for combining interval inputs
--inverted-read-filter -XRF		Inverted (with flipped acceptance/failure conditions) read filters applied before analysis (after regular read filters).
--lenient -LE	false	Lenient processing of VCF files
--max-variants-per-shard	0	If non-zero, partitions VCF output into shards, each containing up to the given number of records.
--QUIET	false	Whether to suppress job-summary info on System.err.
--read-filter -RF		Read filters to be applied before analysis
--read-index		Indices to use for the read inputs. If specified, an index must be provided for every read input and in the same order as the read inputs. If this argument is not specified, the path to the index for each input will be inferred automatically.
--read-validation-stringency -VS	SILENT	Validation stringency for all SAM/BAM/CRAM/SRA files read by this program. The default stringency value SILENT can improve performance when processing a BAM file in which variable-length data (read, qualities, tags) do not otherwise need to be decoded.
--seconds-between-progress-updates	10.0	Output traversal statistics every time this many seconds elapse
--sequence-dictionary		Use the given sequence dictionary as the master/canonical sequence dictionary. Must be a .dict file.
--tmp-dir		Temp directory to use.
--use-jdk-deflater -jdk-deflater	false	Whether to use the JdkDeflater (as opposed to IntelDeflater)
--use-jdk-inflater -jdk-inflater	false	Whether to use the JdkInflater (as opposed to IntelInflater)
--verbosity	INFO	Control verbosity of logging.
Advanced Arguments
--showHidden	false	display hidden arguments

Argument details

Arguments in this list are specific to this tool. Keep in mind that other arguments are available that are shared with other tools (e.g. command-line GATK arguments); see Inherited arguments above.

--add-output-sam-program-record / -add-output-sam-program-record

If true, adds a PG tag to created SAM/BAM/CRAM files.

boolean true

--add-output-vcf-command-line / -add-output-vcf-command-line

If true, adds a command line header line to created VCF files.

boolean true

--allosomal-contig

Contigs to treat as allosomal (i.e. choose their reference copy-number allele according to the sample karyotype).

List[String] []

--arguments_file

read one or more arguments files and add them to the command line

List[File] []

--autosomal-ref-copy-number

Reference copy-number on autosomal intervals.

int 2 [ [ 0 ∞ ] ]

--calls-shard-path

List of paths to GermlineCNVCaller call directories.

R List[File] []

--cloud-index-prefetch-buffer / -CIPB

Size of the cloud-only prefetch buffer (in MB; 0 to disable). Defaults to cloudPrefetchBuffer if unset.

int -1 [ [ -∞ ∞ ] ]

--cloud-prefetch-buffer / -CPB

Size of the cloud-only prefetch buffer (in MB; 0 to disable).

int 40 [ [ -∞ ∞ ] ]

--clustered-breakpoints

VCF with clustered breakpoints and copy number calls for all samples, can be generated with GATK JointGermlineCNVSegmentation tool

File null

--contig-ploidy-calls

Path to contig-ploidy calls directory (output of DetermineGermlineContigPloidy).

R File null

--create-output-bam-index / -OBI

If true, create a BAM/CRAM index when writing a coordinate-sorted BAM/CRAM file.

boolean true

--create-output-bam-md5 / -OBM

If true, create a MD5 digest for any BAM/SAM/CRAM file created

boolean false

--create-output-variant-index / -OVI

If true, create a VCF index when writing a coordinate-sorted VCF file.

boolean true

--create-output-variant-md5 / -OVM

If true, create a a MD5 digest any VCF file created.

boolean false

--disable-bam-index-caching / -DBIC

If true, don't cache bam indexes, this will reduce memory requirements but may harm performance if many intervals are specified. Caching is automatically disabled if there are no intervals specified.

boolean false

--disable-read-filter / -DF

Read filters to be disabled before analysis

List[String] []

--disable-sequence-dictionary-validation / -disable-sequence-dictionary-validation

If specified, do not check the sequence dictionaries from our inputs for compatibility. Use at your own risk!

boolean false

--disable-tool-default-read-filters / -disable-tool-default-read-filters

Disable all tool default read filters (WARNING: many tools will not function correctly without their default read filters on)

boolean false

--duplication-qs-threshold

Filter out duplications with quality lower than this.

int 50 [ [ 0 ∞ ] ]

--exclude-intervals / -XL

One or more genomic intervals to exclude from processing
Use this argument to exclude certain parts of the genome from the analysis (like -L, but the opposite). This argument can be specified multiple times. You can use samtools-style intervals either explicitly on the command line (e.g. -XL 1 or -XL 1:100-200) or by loading in a file containing a list of intervals (e.g. -XL myFile.intervals). strings gathered from the command line -XL argument to be parsed into intervals to exclude

List[String] []

--gatk-config-file

A configuration file to use with the GATK.

String null

--gcs-max-retries / -gcs-retries

If the GCS bucket channel errors out, how many times it will attempt to re-initiate the connection

int 20 [ [ -∞ ∞ ] ]

--gcs-project-for-requester-pays

Project to bill when accessing "requester pays" buckets. If unset, these buckets cannot be accessed. User must have storage.buckets.get permission on the bucket being accessed.

String ""

--help / -h

display the help message

boolean false

--het-deletion-qs-threshold

Filter out heterozygous deletions with quality lower than this.

int 100 [ [ 0 ∞ ] ]

--hom-deletion-qs-threshold

Filter out homozygous deletions with quality lower than this.

int 400 [ [ 0 ∞ ] ]

--input / -I

BAM/SAM/CRAM file containing reads

List[GATKPath] []

--input-intervals-vcf

Input VCF with combined intervals for all samples

File null

--interval-exclusion-padding / -ixp

Amount of padding (in bp) to add to each interval you are excluding.
Use this to add padding to the intervals specified using -XL. For example, '-XL 1:100' with a padding value of 20 would turn into '-XL 1:80-120'. This is typically used to add padding around targets when analyzing exomes.

int 0 [ [ -∞ ∞ ] ]

--interval-merging-rule / -imr

Interval merging rule for abutting intervals
By default, the program merges abutting intervals (i.e. intervals that are directly side-by-side but do not actually overlap) into a single continuous interval. However you can change this behavior if you want them to be treated as separate intervals instead.

The --interval-merging-rule argument is an enumerated type (IntervalMergingRule), which can have one of the following values:

ALL
OVERLAPPING_ONLY

IntervalMergingRule ALL

--interval-padding / -ip

Amount of padding (in bp) to add to each interval you are including.
Use this to add padding to the intervals specified using -L. For example, '-L 1:100' with a padding value of 20 would turn into '-L 1:80-120'. This is typically used to add padding around targets when analyzing exomes.

int 0 [ [ -∞ ∞ ] ]

--interval-set-rule / -isr

Set merging approach to use for combining interval inputs
By default, the program will take the UNION of all intervals specified using -L and/or -XL. However, you can change this setting for -L, for example if you want to take the INTERSECTION of the sets instead. E.g. to perform the analysis only on chromosome 1 exomes, you could specify -L exomes.intervals -L 1 --interval-set-rule INTERSECTION. However, it is not possible to modify the merging approach for intervals passed using -XL (they will always be merged using UNION). Note that if you specify both -L and -XL, the -XL interval set will be subtracted from the -L interval set.

The --interval-set-rule argument is an enumerated type (IntervalSetRule), which can have one of the following values:

UNION: Take the union of all intervals
INTERSECTION: Take the intersection of intervals (the subset that overlaps all intervals specified)

IntervalSetRule UNION

--intervals / -L

One or more genomic intervals over which to operate

List[String] []

--inverted-read-filter / -XRF

Inverted (with flipped acceptance/failure conditions) read filters applied before analysis (after regular read filters).

List[String] []

--lenient / -LE

Lenient processing of VCF files

boolean false

--max-variants-per-shard

If non-zero, partitions VCF output into shards, each containing up to the given number of records.

int 0 [ [ 0 ∞ ] ]

--model-shard-path

List of paths to GermlineCNVCaller model directories.

R List[File] []

--output-denoised-copy-ratios

Output denoised copy ratio file.

R File null

--output-genotyped-intervals

Output intervals VCF file.

R File null

--output-genotyped-segments

Output segments VCF file.

R File null

--QUIET

Whether to suppress job-summary info on System.err.

Boolean false

--read-filter / -RF

Read filters to be applied before analysis

List[String] []

--read-index / -read-index

Indices to use for the read inputs. If specified, an index must be provided for every read input and in the same order as the read inputs. If this argument is not specified, the path to the index for each input will be inferred automatically.

List[GATKPath] []

--read-validation-stringency / -VS

Validation stringency for all SAM/BAM/CRAM/SRA files read by this program. The default stringency value SILENT can improve performance when processing a BAM file in which variable-length data (read, qualities, tags) do not otherwise need to be decoded.

The --read-validation-stringency argument is an enumerated type (ValidationStringency), which can have one of the following values: