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VCF format for complex structural variations

Introduction

VarSim follows the latest VCF format specification 4.3.

Details

  • In VCF Specification 4.3, the only values allowed for SVTYPE in INFO are DEL, INS, DUP, INV, CNV and BND.
  • In ALT column, a subtype can be specified, e.g. <DUP:TANDEM>.
  • In ALT column, values allowed for first level symbolic alleles are DEL, INS, DUP, INV, CNV. Reserved subtypes include DUP:TANDEM, DEL:ME, INS:ME.
  • A cut-and-paste translocation, or simply as translocation, is represented in 2 lines, one as <DEL:TRA>, the other as <DUP:TRA>. That is, they must appear in pairs. Evaluation will be performed independently on these two lines. The two lines can be reported as a single event (translocation) or 2 unrelated events (deletion + duplication). If reported as a single event, VarSim uses TRAID in INFO field to differentiate between different translocations.
  • For a duplication (copy-and-paste), it comes with additional INFO fields (CHR2, POS2, END2) to indicate source of duplicated sequences. If these additional fields are absent, we assume it is tandem duplication otherwise <DUP> and <DUP:ISP> are essentially the same. The first level symbolic allele in ALT must be DUP to indicate that this line denotes a duplication. SVLEN will be used to determine length of duplication regardless of CHR2, POS2 and END2.
  • An additional tag is required in INFO field to indicate the direction of inserted duplicate. VarSim recognizes ISINV. If it is absent, then we assume it is on positive strand.
  • Some programs may report translocations in a different way, their output will be converted to VarSim format before complex variants can be analyzed.
  • Subtypes for DUP may include TRA, TANDEM and ISP (interspersed). Other subtypes are ignored (treated as if they were not there).
  • INS must represent novel sequence insertion (i.e., cannot be duplication). VarSim takes this as a truth without enforcing it.

Examples

Header

The following fields are honored by VarSim. Other fields may be present, but will be ignored. Note here the type of CN is String not Integer. Its format is similar to GT.

##fileformat=VCFv4.3
##reference=referenceFileName.fasta
##INFO=<ID=SVLEN,Number=.,Type=Integer,Description="Difference in length between REF and ALT alleles">
##INFO=<ID=SVTYPE,Number=1,Type=String,Description="Type of structural variant">
##INFO=<ID=CHR2,Number=1,Type=String,Description="Chromosome of source sequence">
##INFO=<ID=POS2,Number=1,Type=Integer,Description="1-based start position of source sequence">
##INFO=<ID=END2,Number=1,Type=Integer,Description="1-based end position of source sequence">
##INFO=<ID=ISINV,Number=1,Type=Flag,Description="whether a duplication is inverted">
##INFO=<ID=TRAID,Number=1,Type=String,Description="translocation ID">
##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">
##FORMAT=<ID=CN,Number=1,Type=String,Description="Copy number for each genotype">
##ALT=<ID=DEL,Description="Deletion">
##ALT=<ID=DEL:TRA,Description="Deletion in translocation">
##ALT=<ID=DUP,Description="Duplication">
##ALT=<ID=DUP:TANDEM,Description="Tandem Duplication">
##ALT=<ID=DUP:ISP,Description="Interspersed duplication">
##ALT=<ID=DUP:TRA,Description="Duplication in translocation">
##ALT=<ID=INS,Description="Insertion of novel sequence">
##ALT=<ID=INV,Description="Inversion">
#CHROM	POS	ID	REF	ALT	QUAL	FILTER	INFO	FORMAT  SAMPLEX SAMPLEY ...

Example complex variants

Deletion

1	4	.	C	<DEL>	.	.	SVTYPE=DEL;SVLEN=-3	GT	1|0

It means reference sequence 1:5-7 is deleted.

Tandem duplication

1	3	.	T	<DUP:TANDEM>	.	PASS	SVTYPE=DUP;SVLEN=4	GT	1|1

The following line is equivalent.

1	3	.	T	<DUP>	.	PASS	SVTYPE=DUP;SVLEN=4	GT	1|1

It means 4bp of reference sequence (1:4-7) is copied and inserted right after 1:3. Copy number is assumed to be 1 by default.

Cut-and-paste translocation

1	1	.	A	<DUP:TRA>	.	PASS	SVTYPE=DUP;SVLEN=3;TRAID=1;CHR2=2;POS2=3;END2=5	GT	1/1
2	2	.	T	<DEL:TRA>	.	PASS	SVTYPE=DEL;SVLEN=-3;TRAID=1	GT	1|1

It means 2:3-5 is cut and pasted right after 1:1.

Interspersed duplication

1	1	.	A	<DUP:ISP>	.	PASS	SVTYPE=DUP;SVLEN=3;CHR2=2;POS2=3;END2=5	GT	1/1

The following line is equivalent.

1	1	.	A	<DUP>	.	PASS	SVTYPE=DUP;SVLEN=3;CHR2=2;POS2=3;END2=5	GT	1/1

It means 3bp of reference sequence (2:3-5) is copied and inserted right after 1:1.

Cut-and-paste translocation with inverted duplication

1	1	.	A	<DUP:TRA>	.	PASS	SVTYPE=DUP;SVLEN=3;ISINV;TRAID=1;CHR2=2;POS2=3;END2=5	GT	1/1
2	2	.	T	<DEL:TRA>	.	PASS	SVTYPE=DEL;SVLEN=-3;TRAID=1	GT	1|1

It means 2:3-5 is cut and pasted right after 1:1 in reverse-complement orientation.

Insertion

1	3	.	T	<INS>	.	.	SVTYPE=INS;SVLEN=3	GT	1/1

It means 3 bp of unknown sequence is inserted after 1:3.

Inversion

1	3	.	T	<INV>	.	PASS	SVTYPE=INV;SVLEN=4	GT	0|1

It means 4 bp of reference sequence (1:4-7) is inverted.

Implementation details

<DUP:TRA>, <DUP:ISP>

VarSim validates translocation duplication via breakends. Each <DUP:TRA> or <DUP:ISP> is broken into two nonadjacent novel breakends, i.e. the two breakends at the outermost positions of duplication insertion.

validation of breakend

Breakends are validated by two criteria:

  1. the locus, plus/minus wiggle.
  2. the oritentation

It's possible that there are inserted sequences before/after breakends, for now, we ignore them.

<DEL:TRA>

A translocation deletion will be treated as a novel breakend, with another breakend next to it.

<TRAID>

Translocation ID is used to link <DUP:TRA> and <DEL:TRA>, each combination acts as if they were one translocation, and will be valided and reported as a single event. Partial matching will have to exceed overlapRatio to make this translocation match with another. In practice, each linked translocation can be decomposed into 3 breakends, each with one more breakend adjacent to it. The translocation will be considered as valided as long as x/3.0 >= overlapRatio.

Because of the linkage established by TRAID, the number of variants reported as TP in json might differ from that in VCF because each translocation is counted once in statistics but has two underly records in VCF.

The lengths of a translocation are represented by the maximum length of underlying variants.

compare_vcf.py

The new compare_vcf.py scripts wraps VarSim's vcfcompare and RTG's vcfeval. Now we can leverage haplotype-based comparison from vcfeval on top of lenient and flexible comparison by VarSim.

Implementation

  • Run vcfcompare inside VarSim to generate TP, FN and FP.
  • Rescue FN and FP by feeding them into vcfeval
  • Combine the results and update numbers.
  • the following diagram illustrates how it works on a high level compare_vcf.py diagram

Known issues

  • VarSim used to prioritize VCFs in the reverse order of arguments. This issue has been fixed. This issue only affects overlapping variants.

references