From 798a5aced3faf3087de5c5938b830a776da244e3 Mon Sep 17 00:00:00 2001 From: clintval Date: Wed, 22 Nov 2023 14:32:59 -0800 Subject: [PATCH 01/33] Add a description of the toolkit to the README --- .gitignore | 1 + README.md | 28 +++++++++++++++++++++++++--- 2 files changed, 26 insertions(+), 3 deletions(-) diff --git a/.gitignore b/.gitignore index f4a3627..3d84bcb 100644 --- a/.gitignore +++ b/.gitignore @@ -1,3 +1,4 @@ +.java-version .idea .idea_modules out diff --git a/README.md b/README.md index ba576ac..08a964f 100644 --- a/README.md +++ b/README.md @@ -7,15 +7,37 @@ [bioconda-badge-link]: https://img.shields.io/conda/dn/bioconda/fgsv.svg?label=Bioconda [bioconda-link]: http://bioconda.github.io/recipes/fgsv/README.html -[github-badge]: https://github.com/fulcrumgenomics/fgsv/actions/workflows/unittests.yaml/badge.svg +[github-badge]: https://github.com/fulcrumgenomics/fgsv/actions/workflows/unittests.yaml/badge.svg?branch=main [github-link]: https://github.com/fulcrumgenomics/fgsv/actions/workflows/unittests.yaml [scala-badge]: https://img.shields.io/badge/language-scala-c22d40.svg [scala-link]: https://www.scala-lang.org/ [license-badge]: https://img.shields.io/badge/license-MIT-blue.svg [license-link]: https://github.com/fulcrumgenomics/fgsv/blob/main/LICENSE -Tools to find evidence for structural variation. +Tools for calling breakpoints and exploring structural variation. ## Documentation -Documentation can be found in the [docs folder](docs/01_Introduction.md) +Documentation can be found in the [docs folder](docs/01_Introduction.md). + +## Introduction to the `fgsv` Toolkit + +The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a query-grouped BAM file as input and scans through each template one at a time, where a template is the full collection of reads and alignments from a single source molecule. +Primary and supplemental alignments for a template are used to construct a “chain” of aligned sub-segments in a way that is order and strand-aware. +These aligned sub-segments relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference genome and importantly, jumps between reference sequences (chromosomes). + +For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater and the minimum inter-read distance per pair must be 1000bp (by default) or greater. +In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored. +At locations where these jumps occur, breakpoints are marked, and the breakpoints are given a unique ID based on the position of the breakpoint, the directionality of the left and right strands, and the other location the aligned sub-segment jumps to. +The output of this process is simply a pileup of candidate breakpoint locations. +The output of this tool is a metrics file tabulating the breakpoints and a BAM file with each alignment having custom tags that indicate which breakpoint the alignment supports (by breakpoint ID), if any. + +Because of variability in short-read sequence data and their alignments, evidence for a single breakpoint may span a few loci near the true breakpoint. +The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one call if they appear to belong to one breakpoint. +This polishing step preserves true positive breakpoint calls and should reduce the number of false positive breakpoint calls. +Adjacent breakpoints are only merged if their left sides map to the same reference sequence, their right rides sides map to the same reference sequence, the strandedness of the left and right aligned sub-segments is the same, and their left and right positions are both within a given length threshold. +One shortcoming of the existing behavior, that should be corrected at some point, is that inter-read breakpoint evidence is considered similarly to inter-pair breakpoint evidence even though inter-read breakpoint evidence often has nucleotide-level alignment resolution and inter-pair breakpoint evidence does not. +The output of this tool is a metrics file tabulating the coalesced breakpoints with all previous breakpoint IDs listed for the new breakpoint call and an estimation of the allele frequency of the call based on the alignments that support the breakpoint. + +The `fgsv` tools are an effective structural variant debugging toolkit but are not meant to be considered as a structural variant calling toolchain in-and-of-itself. +Instead, it’s better to think of the `fgsv` toolkit as an effective “breakpoint caller”. From e3e3dc1faf6a5b96a525d6d433c3970103cb4c01 Mon Sep 17 00:00:00 2001 From: Nobody Date: Wed, 13 Mar 2024 17:31:34 +0000 Subject: [PATCH 02/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 98eeabd..c1cc652 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.1.0-dcaa891. +The following tools are available in fgsv version 0.1.1-798a5ac. ## All tools All tools. From 531a1228960e4c61210fa40c49960e7cd2025114 Mon Sep 17 00:00:00 2001 From: Nobody Date: Fri, 10 May 2024 19:31:11 +0000 Subject: [PATCH 03/33] Generate docs files --- docs/04_Metrics.md | 12 ++++++------ docs/tools/AggregateSvPileup.md | 2 +- docs/tools/SvPileup.md | 2 +- docs/tools/index.md | 3 +-- 4 files changed, 9 insertions(+), 10 deletions(-) diff --git a/docs/04_Metrics.md b/docs/04_Metrics.md index 1cd4c78..df3cace 100644 --- a/docs/04_Metrics.md +++ b/docs/04_Metrics.md @@ -29,12 +29,12 @@ Aggregated cluster of breakpoint pileups |id|String|Combined ID retaining the IDs of all constituent breakpoints| |category|BreakpointCategory|Breakpoint category| |left_contig|String|Contig name for left side of breakpoint| -|left_min_pos|Int|Minimum coordinate of left breakends (1-based)| -|left_max_pos|Int|Maximum coordinate of left breakends (1-based)| +|left_min_pos|Int|Minimum coordinate of left breakends (1-based inclusive)| +|left_max_pos|Int|Maximum coordinate of left breakends (1-based inclusive)| |left_strand|Char|Strand at left breakends| |right_contig|String|Contig name for right side of breakpoint| -|right_min_pos|Int|Minimum coordinate of right breakends (1-based)| -|right_max_pos|Int|Maximum coordinate of right breakends (1-based)| +|right_min_pos|Int|Minimum coordinate of right breakends (1-based inclusive)| +|right_max_pos|Int|Maximum coordinate of right breakends (1-based inclusive)| |right_strand|Char|Strand at right breakends| |split_reads|Int|Total number of split reads supporting the breakpoints in the cluster| |read_pairs|Int|Total number of read pairs supporting the breakpoints in the cluster| @@ -82,10 +82,10 @@ the only information comes from read-pairs and the breakpoint information should |------|----|-----------| |id|String|An ID assigned to the breakpoint that can be used to lookup supporting reads in the BAM.| |left_contig|String|The contig of chromosome on which the left hand side of the breakpoint exists.| -|left_pos|Int|The position (possibly imprecise) of the left-hand breakend (1-based).| +|left_pos|Int|The position (possibly imprecise) of the left-hand breakend (1-based, inclusive).| |left_strand|Char|The strand of the left-hand breakend; sequence reads would traverse this strand in order to arrive at the breakend and transit into the right-hand side of the breakpoint.| |right_contig|String|The contig of chromosome on which the left hand side of the breakpoint exists.| -|right_pos|Int|The position (possibly imprecise) of the right-hand breakend (1-based).| +|right_pos|Int|The position (possibly imprecise) of the right-hand breakend (1-based, inclusive).| |right_strand|Char|The strand of the right-hand breakend;. sequence reads would continue reading onto this strand after transiting the breakpoint from the left breakend| |split_reads|Int|The number of templates/inserts with split-read alignments that identified this breakpoint.| |read_pairs|Int|The number of templates/inserts with read-pair alignments (and without split-read alignments) that identified this breakpoint.| diff --git a/docs/tools/AggregateSvPileup.md b/docs/tools/AggregateSvPileup.md index 012d262..d3e3029 100644 --- a/docs/tools/AggregateSvPileup.md +++ b/docs/tools/AggregateSvPileup.md @@ -36,7 +36,7 @@ of the overlapping target regions are copied from the `SvPiluep` input (if prese The output file is a tab-delimited table with one record per aggregated cluster of pileups. Aggregated pileups are reported with the minimum and maximum (inclusive) coordinates of all pileups in the cluster, a possible putative structural variant event type supported by the pileups, and the sum of read support from all -pileups in the cluster. Positions in this file are 1-based positions. +pileups in the cluster. Positions in this file are 1-based inclusive positions. ## Arguments diff --git a/docs/tools/SvPileup.md b/docs/tools/SvPileup.md index a6290ec..cca2077 100644 --- a/docs/tools/SvPileup.md +++ b/docs/tools/SvPileup.md @@ -15,7 +15,7 @@ Two output files will be created: 1. `.txt`: a tab-delimited file describing SV pileups, one line per breakpoint event. The returned breakpoint will be canonicalized such that the "left" side of the breakpoint will have the lower (or equal to) - position on the genome vs. the "right"s side. Positions in this file are 1-based positions. + position on the genome vs. the "right"s side. Positions in this file are 1-based inclusive positions. 2. `.bam`: a SAM/BAM file containing reads that contain SV breakpoint evidence annotated with SAM tag. diff --git a/docs/tools/index.md b/docs/tools/index.md index f1bc987..f7af0c0 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,8 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-d603e95. - +The following tools are available in fgsv version 0.2.0-0ade11f. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 17e55e4b17346cbd9c00d5eb49f8ab8c674bebbf Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:09:08 -0700 Subject: [PATCH 04/33] Fix up README after a review --- README.md | 83 +++++++++++++++++++++++++++++++--------- docs/img/fgsv-bedpe.png | Bin 0 -> 563167 bytes 2 files changed, 64 insertions(+), 19 deletions(-) create mode 100644 docs/img/fgsv-bedpe.png diff --git a/README.md b/README.md index 12b5bdd..2f3d5a0 100644 --- a/README.md +++ b/README.md @@ -17,7 +17,7 @@ [doi-badge]: https://zenodo.org/badge/454071954.svg [doi-link]: https://zenodo.org/doi/10.5281/zenodo.10452647 -Tools for calling breakpoints and exploring structural variation. +Tools to gather evidence for structural variation via breakpoint detection. ## Documentation @@ -25,22 +25,67 @@ Documentation can be found in the [docs folder](docs/01_Introduction.md). ## Introduction to the `fgsv` Toolkit +The `fgsv` tools are an effective structural variant debugging toolkit but are not meant to be used as a structural variant calling toolchain in-and-of-itself. +Instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. + +When describing structural variation, we use the term breakpoint to mean a junction between two loci and the term breakend to refer to one of the loci in a breakpoint. +Importantly, all point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence. + +### `fgsv SvPileup` + +Collates a pileup of putative structural variant supporting reads. + +```console +❯ fgsv SvPileup \ + --input sample.bam \ + --output sample.svpileup +``` + The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a query-grouped BAM file as input and scans through each template one at a time, where a template is the full collection of reads and alignments from a single source molecule. -Primary and supplemental alignments for a template are used to construct a “chain” of aligned sub-segments in a way that is order and strand-aware. -These aligned sub-segments relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference genome and importantly, jumps between reference sequences (chromosomes). - -For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater and the minimum inter-read distance per pair must be 1000bp (by default) or greater. -In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored. -At locations where these jumps occur, breakpoints are marked, and the breakpoints are given a unique ID based on the position of the breakpoint, the directionality of the left and right strands, and the other location the aligned sub-segment jumps to. -The output of this process is simply a pileup of candidate breakpoint locations. -The output of this tool is a metrics file tabulating the breakpoints and a BAM file with each alignment having custom tags that indicate which breakpoint the alignment supports (by breakpoint ID), if any. - -Because of variability in short-read sequence data and their alignments, evidence for a single breakpoint may span a few loci near the true breakpoint. -The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one call if they appear to belong to one breakpoint. -This polishing step preserves true positive breakpoint calls and should reduce the number of false positive breakpoint calls. -Adjacent breakpoints are only merged if their left sides map to the same reference sequence, their right rides sides map to the same reference sequence, the strandedness of the left and right aligned sub-segments is the same, and their left and right positions are both within a given length threshold. -One shortcoming of the existing behavior, that should be corrected at some point, is that inter-read breakpoint evidence is considered similarly to inter-pair breakpoint evidence even though inter-read breakpoint evidence often has nucleotide-level alignment resolution and inter-pair breakpoint evidence does not. -The output of this tool is a metrics file tabulating the coalesced breakpoints with all previous breakpoint IDs listed for the new breakpoint call and an estimation of the allele frequency of the call based on the alignments that support the breakpoint. - -The `fgsv` tools are an effective structural variant debugging toolkit but are not meant to be considered as a structural variant calling toolchain in-and-of-itself. -Instead, it’s better to think of the `fgsv` toolkit as an effective “breakpoint caller”. +For example, a paired-end read may have an alignment per read: one alignment for read 1 and another alignment for read 2. + +Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the logical ordering of sub-segments and their strandeness in relation to the reference sequence. +These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between chromosomes or contigs. + +For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater, and the minimum inter-read distance across reads (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. +In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. +At locations where these jumps occur, breakpoints are marked and the breakpoints are given a unique ID based on the positions of the breakends and the directionality of the left and right strands leading into each breakend. + +This process creates a collection of candidate breakpoint locations. +The output of this tool is a metrics file tabulating the breakpoints and a BAM file with each breakpoint-supporting alignment having custom tags that indicate which breakpoint the alignment supports. + +### `fgsv AggregateSvPileup` + +Merges nearby pileups of reads supporting putative breakpoints. + +```console +fgsv AggregateSvPileup \ + --bam sample.bam \ + --input sample.svpileup.txt \ + --output sample.svpileup.aggregate.txt +``` + +Because of variability in typical short-read alignments, evidence for a single breakpoint may span a few loci near the true breakend loci. For example, if the breakpoint only has intra-read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci. + +The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to belong to one true breakpoint. +This polishing step preserves true positive breakpoint events and intends to reduce the number of false positive breakpoint events. + +Adjacent breakpoints are only merged if their left breakends map to the same reference sequence, their right breakends map to the same reference sequence, the strandedness of the left and right aligned sub-segments is the same, and their left and right genomic breakend positions are both within a given length threshold. + +One shortcoming of the existing behavior, which should be corrected at some point, is that intra-read breakpoint evidence is considered similarly to inter-pair breakpoint evidence even though intra-read breakpoint evidence often has nucleotide-level alignment resolution and inter-pair breakpoint evidence does not. + +The output of this tool is a metrics file tabulating the coalesced breakpoints with all previous breakpoint IDs listed for the new breakpoint event and an estimation of the allele frequency of the event based on the alignments that support the breakpoint. + +## `AggregateSvPileupToBedPE` + +Convert the output of `AggregateSvPileup` to BEDPE. + +```console +❯ fgsv AggregateSvPileupToBedPE \ + --input sample.svpileup.aggregate.txt \ + --output sample.svpileup.aggregate.bedpe +``` + +The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format) so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. 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zcLAND9=~lTXDdsh88AoDY>(9+_CMzjlFssH@om5dn2Wv@w>TZB+^rC1 zIkKLYD!HX4%@y2`(F5mG9g8C%lV3F$0rLUy0O;JWM>?>+uM^?OflU#wk_3Ki--5TnEJ9n3p;;OPF_`vR`~6WG8M AhyVZp literal 0 HcmV?d00001 From 983dbb76e4962bfcbc1742f707259b0888b8279c Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:09:52 -0700 Subject: [PATCH 05/33] Fix up README after a review --- README.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/README.md b/README.md index 2f3d5a0..2115285 100644 --- a/README.md +++ b/README.md @@ -36,7 +36,7 @@ Importantly, all point intervals (1-length) reported by this toolkit are 1-based Collates a pileup of putative structural variant supporting reads. ```console -❯ fgsv SvPileup \ +fgsv SvPileup \ --input sample.bam \ --output sample.svpileup ``` @@ -81,7 +81,7 @@ The output of this tool is a metrics file tabulating the coalesced breakpoints w Convert the output of `AggregateSvPileup` to BEDPE. ```console -❯ fgsv AggregateSvPileupToBedPE \ +fgsv AggregateSvPileupToBedPE \ --input sample.svpileup.aggregate.txt \ --output sample.svpileup.aggregate.bedpe ``` From 5a903a05da09288f3842102372f86f3a30184759 Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:10:55 -0700 Subject: [PATCH 06/33] Remove outdated intro in Overview --- docs/01_Introduction.md | 12 ------------ 1 file changed, 12 deletions(-) diff --git a/docs/01_Introduction.md b/docs/01_Introduction.md index 2d6262d..e99c2c0 100644 --- a/docs/01_Introduction.md +++ b/docs/01_Introduction.md @@ -6,15 +6,3 @@ The following sections will help you to get started. * [Contributing](03_Contributing.md) * [Metric Descriptions](04_Metrics.md) * [Tools Descriptions](05_Tools.md) - -## Overview - -`fgsv` contains tools for gathering evidence for structural variants -from aligned reads. The `SvPileup` tool searches for split read mappings -and read pairs that map across breakpoints, emitting verbose information -similar to other "piluep" tools for small variant detection, but in this -case for structural variation detection. The `AggregateSvPileup` attempts -to aggregate information across "nearby" pileups, which is useful as often -the genomic start and end of a breakpoint is not always precise. The tools -aim to be as sensitive as possible to find these evidence, but do neither -perform structural variation calling nor genotyping. From a62b275f53b048b8ab4f7f9c3823acbba5eb57a7 Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:11:42 -0700 Subject: [PATCH 07/33] Fixup a sentence --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index 2115285..cdcdbc6 100644 --- a/README.md +++ b/README.md @@ -25,7 +25,7 @@ Documentation can be found in the [docs folder](docs/01_Introduction.md). ## Introduction to the `fgsv` Toolkit -The `fgsv` tools are an effective structural variant debugging toolkit but are not meant to be used as a structural variant calling toolchain in-and-of-itself. +The `fgsv` toolkit contains tools for effective structural variant debugging but are not meant to be used as a structural variant calling toolchain in-and-of-itself. Instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. When describing structural variation, we use the term breakpoint to mean a junction between two loci and the term breakend to refer to one of the loci in a breakpoint. From a8cc755ea95100030c0290fedcdcef7b4b4935bf Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:12:12 -0700 Subject: [PATCH 08/33] Whitespace --- README.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/README.md b/README.md index cdcdbc6..9dba227 100644 --- a/README.md +++ b/README.md @@ -82,8 +82,8 @@ Convert the output of `AggregateSvPileup` to BEDPE. ```console fgsv AggregateSvPileupToBedPE \ - --input sample.svpileup.aggregate.txt \ - --output sample.svpileup.aggregate.bedpe + --input sample.svpileup.aggregate.txt \ + --output sample.svpileup.aggregate.bedpe ``` The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format) so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. For example: From 9304a02dafd470bac8b374d3167381e3730f7175 Mon Sep 17 00:00:00 2001 From: Nobody Date: Fri, 10 May 2024 20:13:33 +0000 Subject: [PATCH 09/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index f7af0c0..705d189 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-0ade11f. +The following tools are available in fgsv version 0.2.0-a8cc755. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 475c3e9ac776ab84a62e4904906a2a1e5680dc56 Mon Sep 17 00:00:00 2001 From: Nobody Date: Fri, 10 May 2024 20:14:27 +0000 Subject: [PATCH 10/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 705d189..b175ebb 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-a8cc755. +The following tools are available in fgsv version 0.2.0-9304a02. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From c2ca29aa1e5c3d50524a844c97efc3fd8057d688 Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 13:25:04 -0700 Subject: [PATCH 11/33] Remove duplicate .gitignore line --- .gitignore | 1 - 1 file changed, 1 deletion(-) diff --git a/.gitignore b/.gitignore index fd77235..d69facc 100644 --- a/.gitignore +++ b/.gitignore @@ -1,4 +1,3 @@ -.java-version .idea .idea_modules .java-version From 009074696157da7762f642ad0831cb8dcd521ddc Mon Sep 17 00:00:00 2001 From: Nobody Date: Fri, 10 May 2024 20:27:24 +0000 Subject: [PATCH 12/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index b175ebb..9178762 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-9304a02. +The following tools are available in fgsv version 0.2.0-c2ca29a. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 70673114d4c8af05da1c2e92376fe60027f89286 Mon Sep 17 00:00:00 2001 From: clintval Date: Fri, 10 May 2024 14:03:40 -0700 Subject: [PATCH 13/33] Small review fixups --- README.md | 12 ++++++------ .../fulcrumgenomics/sv/tools/AggregateSvPileup.scala | 2 +- .../com/fulcrumgenomics/sv/tools/SvPileup.scala | 2 +- 3 files changed, 8 insertions(+), 8 deletions(-) diff --git a/README.md b/README.md index 9dba227..6f436bb 100644 --- a/README.md +++ b/README.md @@ -28,12 +28,12 @@ Documentation can be found in the [docs folder](docs/01_Introduction.md). The `fgsv` toolkit contains tools for effective structural variant debugging but are not meant to be used as a structural variant calling toolchain in-and-of-itself. Instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. -When describing structural variation, we use the term breakpoint to mean a junction between two loci and the term breakend to refer to one of the loci in a breakpoint. +When describing structural variation, we use the term *breakpoint* to mean a junction between two loci and the term *breakend* to refer to one of the loci in a breakpoint. Importantly, all point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence. ### `fgsv SvPileup` -Collates a pileup of putative structural variant supporting reads. +Collates pileups of reads over breakpoint events. ```console fgsv SvPileup \ @@ -41,10 +41,10 @@ fgsv SvPileup \ --output sample.svpileup ``` -The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a query-grouped BAM file as input and scans through each template one at a time, where a template is the full collection of reads and alignments from a single source molecule. +The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each group of alignments for structural variant evidence. For example, a paired-end read may have an alignment per read: one alignment for read 1 and another alignment for read 2. -Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the logical ordering of sub-segments and their strandeness in relation to the reference sequence. +Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the logical ordering of sub-segments and their strandedness in relation to the reference sequence. These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between chromosomes or contigs. For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater, and the minimum inter-read distance across reads (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. @@ -56,7 +56,7 @@ The output of this tool is a metrics file tabulating the breakpoints and a BAM f ### `fgsv AggregateSvPileup` -Merges nearby pileups of reads supporting putative breakpoints. +Aggregates and merges pileups that are likely to support the same breakpoint. ```console fgsv AggregateSvPileup \ @@ -68,7 +68,7 @@ fgsv AggregateSvPileup \ Because of variability in typical short-read alignments, evidence for a single breakpoint may span a few loci near the true breakend loci. For example, if the breakpoint only has intra-read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci. The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to belong to one true breakpoint. -This polishing step preserves true positive breakpoint events and intends to reduce the number of false positive breakpoint events. +This polishing step preserves true positive breakpoint events and is intended to reduce the number of false positive breakpoint events. Adjacent breakpoints are only merged if their left breakends map to the same reference sequence, their right breakends map to the same reference sequence, the strandedness of the left and right aligned sub-segments is the same, and their left and right genomic breakend positions are both within a given length threshold. diff --git a/src/main/scala/com/fulcrumgenomics/sv/tools/AggregateSvPileup.scala b/src/main/scala/com/fulcrumgenomics/sv/tools/AggregateSvPileup.scala index 01fb081..f8e4531 100644 --- a/src/main/scala/com/fulcrumgenomics/sv/tools/AggregateSvPileup.scala +++ b/src/main/scala/com/fulcrumgenomics/sv/tools/AggregateSvPileup.scala @@ -17,7 +17,7 @@ import scala.collection.mutable @clp(group=ClpGroups.BreakpointAndSv, description= """ - |Merges nearby pileups of reads supporting putative breakpoints. + |Aggregates and merges pileups that are likely to support the same breakpoint. | |Takes as input the file of pileups produced by `SvPileup`. That file contains a list of breakpoints, each |consisting of a chromosome, position and strand for each side of the breakpoint, as well as quantified read support diff --git a/src/main/scala/com/fulcrumgenomics/sv/tools/SvPileup.scala b/src/main/scala/com/fulcrumgenomics/sv/tools/SvPileup.scala index 5b2eeca..f74dd59 100644 --- a/src/main/scala/com/fulcrumgenomics/sv/tools/SvPileup.scala +++ b/src/main/scala/com/fulcrumgenomics/sv/tools/SvPileup.scala @@ -39,7 +39,7 @@ object TargetBedRequirement extends FgBioEnum[TargetBedRequirement] { @clp(group=ClpGroups.BreakpointAndSv, description= """ - |Collates a pileup of putative structural variant supporting reads. + |Collates pileups of reads over breakpoint events. | |## Outputs | From 9a45075e6e6679b991128d51839cb99b9db5d37f Mon Sep 17 00:00:00 2001 From: Nobody Date: Fri, 10 May 2024 21:06:17 +0000 Subject: [PATCH 14/33] Generate docs files --- docs/tools/AggregateSvPileup.md | 2 +- docs/tools/SvPileup.md | 2 +- docs/tools/index.md | 6 +++--- 3 files changed, 5 insertions(+), 5 deletions(-) diff --git a/docs/tools/AggregateSvPileup.md b/docs/tools/AggregateSvPileup.md index d3e3029..ab7e1ef 100644 --- a/docs/tools/AggregateSvPileup.md +++ b/docs/tools/AggregateSvPileup.md @@ -7,7 +7,7 @@ title: AggregateSvPileup ## Overview **Group:** Breakpoint and SV Tools -Merges nearby pileups of reads supporting putative breakpoints. +Aggregates and merges pileups that are likely to support the same breakpoint. Takes as input the file of pileups produced by `SvPileup`. That file contains a list of breakpoints, each consisting of a chromosome, position and strand for each side of the breakpoint, as well as quantified read support diff --git a/docs/tools/SvPileup.md b/docs/tools/SvPileup.md index cca2077..7d194ba 100644 --- a/docs/tools/SvPileup.md +++ b/docs/tools/SvPileup.md @@ -7,7 +7,7 @@ title: SvPileup ## Overview **Group:** Breakpoint and SV Tools -Collates a pileup of putative structural variant supporting reads. +Collates pileups of reads over breakpoint events. ## Outputs diff --git a/docs/tools/index.md b/docs/tools/index.md index 9178762..b2b008f 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,16 +4,16 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-c2ca29a. +The following tools are available in fgsv version 0.2.0-7067311. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. |Tool|Description| |----|-----------| -|[AggregateSvPileup](AggregateSvPileup.md)|Merges nearby pileups of reads supporting putative breakpoints| +|[AggregateSvPileup](AggregateSvPileup.md)|Aggregates and merges pileups that are likely to support the same breakpoint| |[FilterAndMerge](FilterAndMerge.md)|Filters and merges SVPileup output| -|[SvPileup](SvPileup.md)|Collates a pileup of putative structural variant supporting reads| +|[SvPileup](SvPileup.md)|Collates pileups of reads over breakpoint events| ## Utility Tools From af562e668774b3b15e9b010f9d2131b98909a355 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 11:55:49 -0400 Subject: [PATCH 15/33] docs: revise docs based on @msto review --- README.md | 18 +++++++++--------- 1 file changed, 9 insertions(+), 9 deletions(-) diff --git a/README.md b/README.md index 6f436bb..850a6cb 100644 --- a/README.md +++ b/README.md @@ -21,7 +21,7 @@ Tools to gather evidence for structural variation via breakpoint detection. ## Documentation -Documentation can be found in the [docs folder](docs/01_Introduction.md). +More detailed documentation can be found in the [docs folder](docs/01_Introduction.md). ## Introduction to the `fgsv` Toolkit @@ -29,7 +29,7 @@ The `fgsv` toolkit contains tools for effective structural variant debugging but Instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. When describing structural variation, we use the term *breakpoint* to mean a junction between two loci and the term *breakend* to refer to one of the loci in a breakpoint. -Importantly, all point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence. +Importantly, all point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence unless otherwise documented. ### `fgsv SvPileup` @@ -42,10 +42,10 @@ fgsv SvPileup \ ``` The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each group of alignments for structural variant evidence. -For example, a paired-end read may have an alignment per read: one alignment for read 1 and another alignment for read 2. +For a simple example, a paired-end read may have one alignment per read: one alignment for read 1 and another alignment for read 2 mapped to different reference sequences. Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the logical ordering of sub-segments and their strandedness in relation to the reference sequence. -These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between chromosomes or contigs. +These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between contigs or chromosomes. For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater, and the minimum inter-read distance across reads (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. @@ -65,16 +65,16 @@ fgsv AggregateSvPileup \ --output sample.svpileup.aggregate.txt ``` -Because of variability in typical short-read alignments, evidence for a single breakpoint may span a few loci near the true breakend loci. For example, if the breakpoint only has intra-read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci. - -The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to belong to one true breakpoint. +The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to support one true breakpoint. This polishing step preserves true positive breakpoint events and is intended to reduce the number of false positive breakpoint events. -Adjacent breakpoints are only merged if their left breakends map to the same reference sequence, their right breakends map to the same reference sequence, the strandedness of the left and right aligned sub-segments is the same, and their left and right genomic breakend positions are both within a given length threshold. +Because of variability in typical short-read alignments due to somatic mutation, sequencing error, or alignment artifact, evidence for a single breakpoint may span a few loci near the true breakend loci. For a more extreme example, if a true breakpoint only has paired read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci and evidence could be merged within this region. + +Adjacent breakpoints are only merged if their left breakends map to the same strand of the same reference sequence, their right breakends map to the same strand of the same reference sequence, and their left and right genomic breakend positions are both within a given length threshold of 10bp (by default). One shortcoming of the existing behavior, which should be corrected at some point, is that intra-read breakpoint evidence is considered similarly to inter-pair breakpoint evidence even though intra-read breakpoint evidence often has nucleotide-level alignment resolution and inter-pair breakpoint evidence does not. -The output of this tool is a metrics file tabulating the coalesced breakpoints with all previous breakpoint IDs listed for the new breakpoint event and an estimation of the allele frequency of the event based on the alignments that support the breakpoint. +The tool outputs a table of aggregated breakpoints and a modified copy of the input BAM file where each alignment is tagged with the ID of the aggregate breakpoint it supports (if any). ## `AggregateSvPileupToBedPE` From 818d158d30600b9b7763c91ef4bf207d4b11d4d7 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 15:58:29 +0000 Subject: [PATCH 16/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index b2b008f..10fe557 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-7067311. +The following tools are available in fgsv version 0.2.0-af562e6. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From a29d0cab495bfae20fff19d6601b6d75502ff426 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:04:30 -0400 Subject: [PATCH 17/33] docs: small docs fixups for clarity and formatting --- README.md | 22 ++++++++++++++-------- 1 file changed, 14 insertions(+), 8 deletions(-) diff --git a/README.md b/README.md index 850a6cb..f1bcc2e 100644 --- a/README.md +++ b/README.md @@ -25,11 +25,14 @@ More detailed documentation can be found in the [docs folder](docs/01_Introducti ## Introduction to the `fgsv` Toolkit -The `fgsv` toolkit contains tools for effective structural variant debugging but are not meant to be used as a structural variant calling toolchain in-and-of-itself. -Instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. +The `fgsv` toolkit contains tools for effective structural variant debugging. +These tools are not meant to be used as a structural variant calling toolchain in-and-of-itself; instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. -When describing structural variation, we use the term *breakpoint* to mean a junction between two loci and the term *breakend* to refer to one of the loci in a breakpoint. -Importantly, all point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence unless otherwise documented. +> [!NOTE] +> When describing structural variation, we use the term **breakpoint** to mean a junction between two loci and the term **breakend** to refer to one of the loci in a breakpoint. + +> [!IMPORTANT] +> All point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence unless otherwise documented. ### `fgsv SvPileup` @@ -44,12 +47,12 @@ fgsv SvPileup \ The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each group of alignments for structural variant evidence. For a simple example, a paired-end read may have one alignment per read: one alignment for read 1 and another alignment for read 2 mapped to different reference sequences. -Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the logical ordering of sub-segments and their strandedness in relation to the reference sequence. +Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between contigs or chromosomes. For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater, and the minimum inter-read distance across reads (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. -In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. At locations where these jumps occur, breakpoints are marked and the breakpoints are given a unique ID based on the positions of the breakends and the directionality of the left and right strands leading into each breakend. +In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. This process creates a collection of candidate breakpoint locations. The output of this tool is a metrics file tabulating the breakpoints and a BAM file with each breakpoint-supporting alignment having custom tags that indicate which breakpoint the alignment supports. @@ -68,7 +71,9 @@ fgsv AggregateSvPileup \ The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to support one true breakpoint. This polishing step preserves true positive breakpoint events and is intended to reduce the number of false positive breakpoint events. -Because of variability in typical short-read alignments due to somatic mutation, sequencing error, or alignment artifact, evidence for a single breakpoint may span a few loci near the true breakend loci. For a more extreme example, if a true breakpoint only has paired read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci and evidence could be merged within this region. +Because of variability in typical short-read alignments due to somatic mutation, sequencing error, or alignment artifact, evidence for a single breakpoint may span a few loci near the true breakend loci. +For a more extreme example, if a true breakpoint only has paired read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. +In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci and evidence could be merged within this region. Adjacent breakpoints are only merged if their left breakends map to the same strand of the same reference sequence, their right breakends map to the same strand of the same reference sequence, and their left and right genomic breakend positions are both within a given length threshold of 10bp (by default). @@ -86,6 +91,7 @@ fgsv AggregateSvPileupToBedPE \ --output sample.svpileup.aggregate.bedpe ``` -The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format) so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. For example: +The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format) so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. +For example: ![BEDPE in IGV](docs/img/fgsv-bedpe.png) From c6a7e11e37eedcc8211c8a00f5f732976b5f0c9c Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 16:07:06 +0000 Subject: [PATCH 18/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 10fe557..7344f7e 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-af562e6. +The following tools are available in fgsv version 0.2.0-a29d0ca. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 8d80f2a740055dbb2a18275aa61b3355ec598da8 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:21:37 -0400 Subject: [PATCH 19/33] docs: one more pass at docs clarity! --- README.md | 24 ++++++++++++------------ 1 file changed, 12 insertions(+), 12 deletions(-) diff --git a/README.md b/README.md index f1bcc2e..e2916dc 100644 --- a/README.md +++ b/README.md @@ -45,17 +45,17 @@ fgsv SvPileup \ ``` The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each group of alignments for structural variant evidence. -For a simple example, a paired-end read may have one alignment per read: one alignment for read 1 and another alignment for read 2 mapped to different reference sequences. +For a simple example: a paired-end read may have one alignment per read—one alignment for read 1 and another alignment for read 2 mapped to different reference sequences supporting a putative translocation. Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. -These aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences such as translocations between contigs or chromosomes. - -For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment distance within a read must be 100bp (by default) or greater, and the minimum inter-read distance across reads (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. -At locations where these jumps occur, breakpoints are marked and the breakpoints are given a unique ID based on the positions of the breakends and the directionality of the left and right strands leading into each breakend. -In the case where there is both evidence for a split-read alignment and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. +The aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences which could indicate translocations. +For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment jump distance within a read must be 100bp (by default) or greater, and the minimum inter-read jump distance (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. +At locations where these jumps occur, breakpoints are marked and given a unique ID based on the loci of the breakends and the directionality of the left and right strands leading into each breakend. +In the case where there is both evidence for a split-read jump and inter-read jump, the split-read alignment evidence is favored since it gives a precise breakpoint. This process creates a collection of candidate breakpoint locations. -The output of this tool is a metrics file tabulating the breakpoints and a BAM file with each breakpoint-supporting alignment having custom tags that indicate which breakpoint the alignment supports. + +The tool outputs a table of candidate breakpoints and a BAM file with each alignment tagged with the ID of the breakpoint it supports (if any). ### `fgsv AggregateSvPileup` @@ -68,14 +68,14 @@ fgsv AggregateSvPileup \ --output sample.svpileup.aggregate.txt ``` -The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to coalesce nearby breakpoints into one event if they appear to support one true breakpoint. +The tool [`fgsv AggregateSvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileup.md) is used to aggregate nearby breakpoints into one event if they appear to support one true breakpoint. This polishing step preserves true positive breakpoint events and is intended to reduce the number of false positive breakpoint events. -Because of variability in typical short-read alignments due to somatic mutation, sequencing error, or alignment artifact, evidence for a single breakpoint may span a few loci near the true breakend loci. -For a more extreme example, if a true breakpoint only has paired read evidence, then the breakpoint could coincidentally occur within the unobserved bases between read 1 and read 2 in a pair. -In other cases and due to sequence similarity or homology between each breakend locus, it is not always possible to locate the exact nucleotide point where the breakends occur, and instead a plausible region may exist that supports either breakend loci and evidence could be merged within this region. +Aggregating breakpoints is often necessary because of variability in typical short-read alignments caused by somatic mutation, sequencing error, alignment artifact, or breakend sequence similarity/homology to the reference sequence. +Variability in short-read alignments means that it is not always possible to locate the exact nucleotide coordinate where either breakends in a breakpoint occur. +Instead, either breakend of a true breakpoint may map to a plausible region (instead of a point coordinate) and when this happens, the cluster of breakends could be aggregated to build up support for one true breakpoint. -Adjacent breakpoints are only merged if their left breakends map to the same strand of the same reference sequence, their right breakends map to the same strand of the same reference sequence, and their left and right genomic breakend positions are both within a given length threshold of 10bp (by default). +Clustered breakpoints are only merged if their left breakends map to the same strand of the same reference sequence, their right breakends map to the same strand of the same reference sequence, and their left and right genomic breakend positions are both within a given length threshold of 10bp (by default). One shortcoming of the existing behavior, which should be corrected at some point, is that intra-read breakpoint evidence is considered similarly to inter-pair breakpoint evidence even though intra-read breakpoint evidence often has nucleotide-level alignment resolution and inter-pair breakpoint evidence does not. From 88fff290fe4a0ee3a28f9bed69f883977e110706 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:23:29 -0400 Subject: [PATCH 20/33] chore: query group and template definition --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index e2916dc..7769d8f 100644 --- a/README.md +++ b/README.md @@ -44,7 +44,7 @@ fgsv SvPileup \ --output sample.svpileup ``` -The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each group of alignments for structural variant evidence. +The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each query group (template) of alignments for structural variant evidence. For a simple example: a paired-end read may have one alignment per read—one alignment for read 1 and another alignment for read 2 mapped to different reference sequences supporting a putative translocation. Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. From 697b07df35bd797d40abc588b6350ef1bc69f396 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:24:29 -0400 Subject: [PATCH 21/33] docs: move reference down --- README.md | 3 ++- 1 file changed, 2 insertions(+), 1 deletion(-) diff --git a/README.md b/README.md index 7769d8f..87b49c6 100644 --- a/README.md +++ b/README.md @@ -47,8 +47,9 @@ fgsv SvPileup \ The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each query group (template) of alignments for structural variant evidence. For a simple example: a paired-end read may have one alignment per read—one alignment for read 1 and another alignment for read 2 mapped to different reference sequences supporting a putative translocation. -Primary and supplementary alignments for a template (see the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information) are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. +Primary and supplementary alignments for a template are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. The aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences which could indicate translocations. +See the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information on how reads relate to alignments. For each chain of aligned sub-segments per template, outlier jumps are collected where the minimum inter-segment jump distance within a read must be 100bp (by default) or greater, and the minimum inter-read jump distance (e.g. between reads in a paired-end read) must be 1000bp (by default) or greater. At locations where these jumps occur, breakpoints are marked and given a unique ID based on the loci of the breakends and the directionality of the left and right strands leading into each breakend. From 34e9fafd841f7bdf96aed08e674f4b1bdb96a236 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:25:15 -0400 Subject: [PATCH 22/33] docs: do not repeat thyself --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index 87b49c6..f9f381f 100644 --- a/README.md +++ b/README.md @@ -47,7 +47,7 @@ fgsv SvPileup \ The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each query group (template) of alignments for structural variant evidence. For a simple example: a paired-end read may have one alignment per read—one alignment for read 1 and another alignment for read 2 mapped to different reference sequences supporting a putative translocation. -Primary and supplementary alignments for a template are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness in relation to the reference sequence. +Primary and supplementary alignments for a template are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness as compared to the reference sequence. The aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences which could indicate translocations. See the [SAM Format Specification v1](https://samtools.github.io/hts-specs/SAMv1.pdf) for more information on how reads relate to alignments. From ecf1df4c792bf4acbe01f5b35843768d099d4498 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 16:27:28 +0000 Subject: [PATCH 23/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 7344f7e..64f4122 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-a29d0ca. +The following tools are available in fgsv version 0.2.0-34e9faf. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 09ead37867e81e47f28686eb949c131597953921 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 12:27:44 -0400 Subject: [PATCH 24/33] docs: little fixup --- README.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/README.md b/README.md index f9f381f..01d980c 100644 --- a/README.md +++ b/README.md @@ -84,7 +84,7 @@ The tool outputs a table of aggregated breakpoints and a modified copy of the in ## `AggregateSvPileupToBedPE` -Convert the output of `AggregateSvPileup` to BEDPE. +Converts the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format). ```console fgsv AggregateSvPileupToBedPE \ @@ -92,7 +92,7 @@ fgsv AggregateSvPileupToBedPE \ --output sample.svpileup.aggregate.bedpe ``` -The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format) so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. +The tool [`fgsv AggregateSvPileupToBedPE`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/AggregateSvPileupToBedPE.md) is used to convert the output of `AggregateSvPileup` to BEDPE so that it can be viewed in [IGV](https://igv.org/) and other BEDPE-supporting genome browsers. For example: ![BEDPE in IGV](docs/img/fgsv-bedpe.png) From 45a17c11f3b1b86b43bb9184e8342be62a8d2b95 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 16:30:35 +0000 Subject: [PATCH 25/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 64f4122..5f33a8f 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-34e9faf. +The following tools are available in fgsv version 0.2.0-09ead37. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From ef7f8f2df6ed893ce1eaaacdb9ad48e4b1732569 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 13:04:24 -0400 Subject: [PATCH 26/33] docs: formatting to be the same --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index 01d980c..05d6426 100644 --- a/README.md +++ b/README.md @@ -82,7 +82,7 @@ One shortcoming of the existing behavior, which should be corrected at some poin The tool outputs a table of aggregated breakpoints and a modified copy of the input BAM file where each alignment is tagged with the ID of the aggregate breakpoint it supports (if any). -## `AggregateSvPileupToBedPE` +## `fgsv AggregateSvPileupToBedPE` Converts the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format). From ac5b3341a2fd12f795b3c459910f9f7a3536ac41 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 17:07:13 +0000 Subject: [PATCH 27/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 5f33a8f..314e36c 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-09ead37. +The following tools are available in fgsv version 0.2.0-ef7f8f2. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From f56f0e7dec8996a3528fc38734542b304fce55f9 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 13:21:47 -0400 Subject: [PATCH 28/33] chore: header fixup --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index 05d6426..a83b8dd 100644 --- a/README.md +++ b/README.md @@ -82,7 +82,7 @@ One shortcoming of the existing behavior, which should be corrected at some poin The tool outputs a table of aggregated breakpoints and a modified copy of the input BAM file where each alignment is tagged with the ID of the aggregate breakpoint it supports (if any). -## `fgsv AggregateSvPileupToBedPE` +### `fgsv AggregateSvPileupToBedPE` Converts the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format). From 229732451899f3bbcd717fc3879423d6ceed0ef0 Mon Sep 17 00:00:00 2001 From: clintval Date: Tue, 23 Jul 2024 13:22:41 -0400 Subject: [PATCH 29/33] chore: header fixup --- README.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/README.md b/README.md index a83b8dd..b46d77a 100644 --- a/README.md +++ b/README.md @@ -34,7 +34,7 @@ These tools are not meant to be used as a structural variant calling toolchain i > [!IMPORTANT] > All point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence unless otherwise documented. -### `fgsv SvPileup` +### `SvPileup` Collates pileups of reads over breakpoint events. @@ -58,7 +58,7 @@ This process creates a collection of candidate breakpoint locations. The tool outputs a table of candidate breakpoints and a BAM file with each alignment tagged with the ID of the breakpoint it supports (if any). -### `fgsv AggregateSvPileup` +### `AggregateSvPileup` Aggregates and merges pileups that are likely to support the same breakpoint. @@ -82,7 +82,7 @@ One shortcoming of the existing behavior, which should be corrected at some poin The tool outputs a table of aggregated breakpoints and a modified copy of the input BAM file where each alignment is tagged with the ID of the aggregate breakpoint it supports (if any). -### `fgsv AggregateSvPileupToBedPE` +### `AggregateSvPileupToBedPE` Converts the output of `AggregateSvPileup` to the [BEDPE format](https://bedtools.readthedocs.io/en/latest/content/general-usage.html#bedpe-format). From 51557f2fb0a46c95afae2eb6ed14c5753d6dad04 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 17:24:32 +0000 Subject: [PATCH 30/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 314e36c..21ddf60 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-ef7f8f2. +The following tools are available in fgsv version 0.2.0-2297324. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From df8ead11a492d35a78a6d9043fa6673876c2abe7 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 23 Jul 2024 17:25:37 +0000 Subject: [PATCH 31/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 21ddf60..10fc508 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-2297324. +The following tools are available in fgsv version 0.2.0-51557f2. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs. From 3f52590b0dd2752305ef089fa1ec40cbedc828e3 Mon Sep 17 00:00:00 2001 From: clintval Date: Mon, 12 Aug 2024 18:54:20 -0700 Subject: [PATCH 32/33] docs: suit review from @nh13 --- README.md | 8 ++++---- 1 file changed, 4 insertions(+), 4 deletions(-) diff --git a/README.md b/README.md index b46d77a..6b5a88e 100644 --- a/README.md +++ b/README.md @@ -25,11 +25,11 @@ More detailed documentation can be found in the [docs folder](docs/01_Introducti ## Introduction to the `fgsv` Toolkit -The `fgsv` toolkit contains tools for effective structural variant debugging. -These tools are not meant to be used as a structural variant calling toolchain in-and-of-itself; instead, it is better to think of `fgsv` as an effective breakpoint detection and structural variant exploration toolkit. +The `fgsv` toolkit contains tools for effective structural variant investigation. +These tools are not meant to be used as a structural variant calling toolchain in-and-of-itself; instead, it is better to think of `fgsv` as a breakpoint detection and structural variant exploration toolkit. > [!NOTE] -> When describing structural variation, we use the term **breakpoint** to mean a junction between two loci and the term **breakend** to refer to one of the loci in a breakpoint. +> When describing structural variation, we use the term **breakpoint** to mean a junction between two loci and the term **breakend** to refer to one of the loci on one side of a breakpoint. > [!IMPORTANT] > All point intervals (1-length) reported by this toolkit are 1-based inclusive from the perspective of the reference sequence unless otherwise documented. @@ -45,7 +45,7 @@ fgsv SvPileup \ ``` The tool [`fgsv SvPileup`](https://github.com/fulcrumgenomics/fgsv/blob/main/docs/tools/SvPileup.md) takes a queryname-grouped BAM file as input and scans each query group (template) of alignments for structural variant evidence. -For a simple example: a paired-end read may have one alignment per read—one alignment for read 1 and another alignment for read 2 mapped to different reference sequences supporting a putative translocation. +For a simple example: a paired-end read may have one alignment per read (one alignment for read 1 and another alignment for read 2) mapped to different reference sequences supporting a putative translocation. Primary and supplementary alignments for a template are used to construct a “chain” of aligned sub-segments in a way that honors the sub-segments mapping locations and strandedness as compared to the reference sequence. The aligned sub-segments in a chain relate to each other through typical alignment mechanisms like insertions and deletions but also contain information about the relative orientation of the sub-segment to the reference sequence and importantly, jumps between reference sequences which could indicate translocations. From ddd45aab94e653923b3cc987d11d8e9c7191b4e1 Mon Sep 17 00:00:00 2001 From: Nobody Date: Tue, 13 Aug 2024 01:57:16 +0000 Subject: [PATCH 33/33] Generate docs files --- docs/tools/index.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/tools/index.md b/docs/tools/index.md index 10fc508..0c8fe57 100644 --- a/docs/tools/index.md +++ b/docs/tools/index.md @@ -4,7 +4,7 @@ title: fgsv tools # fgsv tools -The following tools are available in fgsv version 0.2.0-51557f2. +The following tools are available in fgsv version 0.2.0-3f52590. ## Breakpoint and SV Tools Primary tools for calling and transforming breakpoints and SVs.