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VCFv4.3.tex
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\documentclass[8pt]{article}
\usepackage{enumerate}
\usepackage{graphicx}
\usepackage{tabularx}
\usepackage{lscape}
\usepackage[margin=0.75in]{geometry}
\usepackage[pdfborder={0 0 0}]{hyperref}
\usepackage{listings}
\lstset{
basicstyle=\ttfamily,
mathescape
}
\usepackage{color}
\renewcommand{\thefootnote}{\fnsymbol{footnote}}
\begin{document}
\input{VCFv4.3.ver}
\title{The Variant Call Format Specification \\ \vspace{0.5em} \large VCFv4.3 and BCFv2.2}
\date{\headdate}
\maketitle
\begin{quote}\small
The master version of this document can be found at
\url{https://github.com/samtools/hts-specs}.\\
This printing is version~\commitdesc\ from that repository,
last modified on the date shown above.
\end{quote}
\vspace*{1em}
\newpage
\tableofcontents
\newpage
\section{The VCF specification}
VCF is a text file format (most likely stored in a compressed manner).
It contains meta-information lines (prefixed with "\#\#"), a header
line (prefixed with "\#"), and data lines
each containing information about a position in the genome and genotype
information on samples for each position
(text fields separated by tabs). Zero length fields are not allowed, a dot (".") must
be used instead.
In order to ensure interoperability across platforms, VCF compliant implementations must support
both LF (\texttt{\textbackslash n}) and CR+LF (\texttt{\textbackslash r\textbackslash n}) newline conventions.
\subsection{An example}
\scriptsize
\begin{verbatim}
##fileformat=VCFv4.3
##fileDate=20090805
##source=myImputationProgramV3.1
##reference=file:///seq/references/1000GenomesPilot-NCBI36.fasta
##contig=<ID=20,length=62435964,assembly=B36,md5=f126cdf8a6e0c7f379d618ff66beb2da,species="Homo sapiens",taxonomy=x>
##phasing=partial
##INFO=<ID=NS,Number=1,Type=Integer,Description="Number of Samples With Data">
##INFO=<ID=DP,Number=1,Type=Integer,Description="Total Depth">
##INFO=<ID=AF,Number=A,Type=Float,Description="Allele Frequency">
##INFO=<ID=AA,Number=1,Type=String,Description="Ancestral Allele">
##INFO=<ID=DB,Number=0,Type=Flag,Description="dbSNP membership, build 129">
##INFO=<ID=H2,Number=0,Type=Flag,Description="HapMap2 membership">
##FILTER=<ID=q10,Description="Quality below 10">
##FILTER=<ID=s50,Description="Less than 50% of samples have data">
##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">
##FORMAT=<ID=GQ,Number=1,Type=Integer,Description="Genotype Quality">
##FORMAT=<ID=DP,Number=1,Type=Integer,Description="Read Depth">
##FORMAT=<ID=HQ,Number=2,Type=Integer,Description="Haplotype Quality">
#CHROM POS ID REF ALT QUAL FILTER INFO FORMAT NA00001 NA00002 NA00003
20 14370 rs6054257 G A 29 PASS NS=3;DP=14;AF=0.5;DB;H2 GT:GQ:DP:HQ 0|0:48:1:51,51 1|0:48:8:51,51 1/1:43:5:.,.
20 17330 . T A 3 q10 NS=3;DP=11;AF=0.017 GT:GQ:DP:HQ 0|0:49:3:58,50 0|1:3:5:65,3 0/0:41:3
20 1110696 rs6040355 A G,T 67 PASS NS=2;DP=10;AF=0.333,0.667;AA=T;DB GT:GQ:DP:HQ 1|2:21:6:23,27 2|1:2:0:18,2 2/2:35:4
20 1230237 . T . 47 PASS NS=3;DP=13;AA=T GT:GQ:DP:HQ 0|0:54:7:56,60 0|0:48:4:51,51 0/0:61:2
20 1234567 microsat1 GTC G,GTCT 50 PASS NS=3;DP=9;AA=G GT:GQ:DP 0/1:35:4 0/2:17:2 1/1:40:3
\end{verbatim}
\normalsize
This example shows (in order): a good simple SNP, a possible SNP that has been filtered out because its quality is below 10, a site at which two alternate alleles are called, with one of them (T) being ancestral (possibly a reference sequencing error), a site that is called monomorphic reference (i.e. with no alternate alleles), and a microsatellite with two alternative alleles, one a deletion of 2 bases (TC), and the other an insertion of one base (T). Genotype data are given for three samples, two of which are phased and the third unphased, with per sample genotype quality, depth and haplotype qualities (the latter only for the phased samples) given as well as the genotypes. The microsatellite calls are unphased.
\subsection{Character encoding, non-printable characters and characters with special meaning}
\label{character-encoding}
The character encoding of VCF files is UTF-8. UTF-8 is a multi-byte
character encoding that is a strict superset of 7-bit ASCII and has the
property that none of the bytes in any multi-byte characters are 7-bit ASCII
bytes. As a result, most software that processes VCF files does not have
to be aware of the possible presence of multi-byte UTF-8 characters.
Note that non-printable characters U+0000-U+0008, U+000B-U+000C, U+000E-U+001F are disallowed.
Line separators must be CR+LF or LF and they are allowed only as line separators at
end of line.
Characters with special meaning (such as field delimiters ';' in INFO
or ':' FORMAT fields) must be represented using the capitalized percent encoding:
\begingroup\footnotesize
\begin{tabular}{l l l}
\%3A & : & (colon) \\
\%3B & ; & (semicolon) \\
\%3D & = & (equal sign) \\
\%25 & \% & (percent sign) \\
\%2C & , & (comma) \\
\%0D & CR & \\
\%0A & LF & \\
\%09 & TAB &
\end{tabular}
\endgroup
\subsection{Data types}
Data types supported by VCF are: Integer (32-bit, signed), Float (32-bit, formatted
to match the regular expression \texttt{\^{}[-+]?[0-9]*\textbackslash.?[0-9]+([eE][-+]?[0-9]+)?\$}, \texttt{NaN}, or \texttt{+/-Inf}), Flag, Character, and
String. For the Integer type, the values from $-2^{31}$ to $-2^{31}+7$ cannot be stored in the binary version and therefore
are disallowed in both VCF and BCF, see \ref{BcfTypeEncoding}.
\subsection{Meta-information lines}
File meta-information is included after the \#\# string and must be key=value
pairs. Meta-information lines are optional, but if they are present then
they must be completely well-formed. Note that BCF, the binary
counterpart of VCF, requires that all entries are present. It is recommended
to include meta-information lines describing the entries used in the
body of the VCF file.
All structured lines that have their value enclosed within "$<>$" require an ID
which must be unique within their type. For all of the structured lines (\#\#INFO, \#\#FORMAT,
\#\#FILTER, etc.), extra fields can be included after the default fields. For example:
\begin{verbatim}
##INFO=<ID=ID,Number=number,Type=type,Description="description",Source="description",Version="128">
\end{verbatim}
In the above example, the extra fields of ``Source'' and ``Version'' are
provided. Optional fields must be stored as strings even for numeric values.
It is recommended in VCF and required in BCF that the header
includes tags describing the reference and contigs backing the data contained in
the file. These tags are based on the SQ field from the SAM spec; all tags are
optional (see the VCF example above).
Meta-information lines can be in any order with the exception of `fileformat`
which must come first.
\subsubsection{File format}
A single `fileformat' line is always required, must be the first line in the file, and details the VCF format version number. For VCF version 4.3, this line is:
\begin{verbatim}
##fileformat=VCFv4.3
\end{verbatim}
\subsubsection{Information field format}
INFO fields are described as follows (first four keys are required, source and version are recommended):
\begin{verbatim}
##INFO=<ID=ID,Number=number,Type=type,Description="description",Source="source",Version="version">
\end{verbatim}
Possible Types for INFO fields are: Integer, Float, Flag, Character, and
String.
The Number entry is an Integer that describes the number of values that
can be included with the INFO field. For example, if the INFO field contains a
single number, then this value must be $1$; if the INFO field describes a
pair of numbers, then this value must be $2$ and so on. There are also
certain special characters used to define special cases:
\begin{itemize}
\item A: The field has one value per alternate allele.
\item R: The field has one value for each possible allele, including the reference.
\item G: The field has one value for each possible genotype (more relevant to the FORMAT tags).
\item . (dot): The number of possible values varies, is unknown or unbounded.
\end{itemize}
The `Flag' type indicates that the INFO field does not contain a Value entry, and hence the Number must be $0$ in this case. The Description value must be surrounded by double-quotes. Double-quote character must be escaped with backslash $\backslash$ and backslash as $\backslash\backslash$. Source and Version values likewise must be surrounded by double-quotes and specify the annotation source (case-insensitive, e.g. ``dbsnp'') and exact version (e.g. ``138''), respectively for computational use.
\subsubsection{Filter field format}
FILTERs that have been applied to the data are described as follows:
\begin{verbatim}
##FILTER=<ID=ID,Description="description">
\end{verbatim}
\subsubsection{Individual format field format}
Genotype fields specified in the FORMAT field are described as follows:
\begin{verbatim}
##FORMAT=<ID=ID,Number=number,Type=type,Description="description">
\end{verbatim}
Possible Types for FORMAT fields are: Integer, Float, Character, and String (this field is otherwise defined precisely as the INFO field).
\subsubsection{Alternative allele field format}
Symbolic alternate alleles are described as follows:
\begin{verbatim}
##ALT=<ID=type,Description=description>
\end{verbatim}
\noindent \textbf{Structural Variants} \newline
In symbolic alternate alleles for imprecise structural variants,
the ID field indicates the type of structural variant, and can be a
colon-separated list of types and subtypes. ID values are case sensitive
strings and must not contain whitespace or angle brackets. The first level type
must be one of the following:
\begin{itemize}
\item DEL Deletion relative to the reference
\item INS Insertion of novel sequence relative to the reference
\item DUP Region of elevated copy number relative to the reference
\item INV Inversion of reference sequence
\item CNV Copy number variable region (may be both deletion and duplication)
\end{itemize}
The CNV category should not be used when a more specific category can be applied. Reserved subtypes include:
\begin{itemize}
\item DUP:TANDEM Tandem duplication
\item DEL:ME Deletion of mobile element relative to the reference
\item INS:ME Insertion of a mobile element relative to the reference
\end{itemize}
\bigskip
\noindent \textbf{IUPAC ambiguity codes} \newline
Symbolic alleles can be used also to represent genuinely ambiguous data in VCF, for example:
\begin{verbatim}
##ALT=<ID=R,Description="IUPAC code R = A/G">
##ALT=<ID=M,Description="IUPAC code M = A/C">
\end{verbatim}
\subsubsection{Assembly field format}
Breakpoint assemblies for structural variations may use an external file:
\begin{verbatim}
##assembly=url
\end{verbatim}
The URL field specifies the location of a fasta file containing breakpoint assemblies referenced in the VCF records for structural variants via the BKPTID INFO key.
\subsubsection{Contig field format}
\label{sec-contig-field}
It is recommended for VCF, and required for BCF, that the header includes tags
describing the contigs referred to in the file. The structured \texttt{contig}
field must include the ID attribute and typically includes also
sequence length, MD5 checksum, URL tag to indicate where the sequence can be
found, etc. For example:
\begin{verbatim}
##contig=<ID=ctg1,length=81195210,URL=ftp://somewhere.org/assembly.fa,...>
\end{verbatim}
\noindent
Valid contig names must follow the reference sequence names allowed by the SAM format
("{\tt [!-)+-\char60\char62-\char126][!-\char126]*}") excluding the characters "\texttt{\textless\textgreater[]:*}" to avoid clashes with
symbolic alleles and breakend notation. The contig names must not use a reserved symbolic allele name.
\subsubsection{Sample field format}
It is possible to define sample to genome mappings as shown below:
{\scriptsize
\begin{verbatim}
##META=<ID=Assay,Type=String,Number=.,Values=[WholeGenome, Exome]>
##META=<ID=Disease,Type=String,Number=.,Values=[None, Cancer]>
##META=<ID=Ethnicity,Type=String,Number=.,Values=[AFR, CEU, ASN, MEX]>
##META=<ID=Tissue,Type=String,Number=.,Values=[Blood, Breast, Colon, Lung, ?]>
##SAMPLE=<ID=Sample1,Assay=WholeGenome,Ethnicity=AFR,Disease=None,Description="Patient germline genome from unaffected",DOI=url>
##SAMPLE=<ID=Sample2,Assay=Exome,Ethnicity=CEU,Disease=Cancer,Tissue=Breast,Description="European patient exome from breast cancer">
\end{verbatim}}
\subsubsection{Pedigree field format}
It is possible to record relationships between genomes using the following syntax:
\begin{verbatim}
##PEDIGREE=<ID=TumourSample,Original=GermlineID>
##PEDIGREE=<ID=SomaticNonTumour,Original=GermlineID>
##PEDIGREE=<ID=ChildID,Father=FatherID,Mother=MotherID>
##PEDIGREE=<ID=SampleID,Name_1=Ancestor_1,...,Name_N=Ancestor_N>
\end{verbatim}
\noindent or a link to a database:
\begin{verbatim}
##pedigreeDB=URL
\end{verbatim}
\noindent See \ref{PedigreeInDetail} for details.
\subsection{Header line syntax}
The header line names the 8 fixed, mandatory columns. These columns are as follows:
\begin{enumerate}
\item \#CHROM
\item POS
\item ID
\item REF
\item ALT
\item QUAL
\item FILTER
\item INFO
\end{enumerate}
If genotype data is present in the file, these are followed by a FORMAT column header, then an arbitrary number of sample IDs. The header line is tab-delimited
and there must be no tab characters at the end of the line.
\subsection{Data lines}
All data lines are tab-delimited
with no tab character at the end of the line. The last data line must end with a line separator. In all cases,
missing values are specified with a dot (`.').
\subsubsection{Fixed fields}
There are 8 fixed fields per record. Fixed fields are:
\begin{enumerate}
\item CHROM - chromosome: An identifier from the reference genome or an angle-bracketed ID String (``$<$ID$>$'') pointing to a contig in the assembly file (cf. the \#\#assembly line in the header). All entries for a specific CHROM must form a contiguous block within the VCF file. The colon symbol (:) must be absent from all chromosome names to avoid parsing errors when dealing with breakends. (String, no white-space permitted, Required).
\item POS - position: The reference position, with the 1st base having position 1. Positions are sorted numerically, in increasing order, within each reference sequence CHROM. It is permitted to have multiple records with the same POS. Telomeres are indicated by using positions 0 or N+1, where N is the length of the corresponding chromosome or contig. (Integer, Required)
\item ID - identifier: Semi-colon separated list of unique identifiers where available. If this is a dbSNP variant the rs number(s) should be used. No identifier should be present in more than one data record. If there is no identifier available, then the missing value should be used. (String, no white-space or semi-colons permitted, duplicate values not allowed.)
\item REF - reference base(s): Each base must be one of A,C,G,T,N (case
insensitive). Multiple bases are permitted. The value in the POS field refers
to the position of the first base in the String. For simple insertions and
deletions in which either the REF or one of the ALT alleles would otherwise
be null/empty, the REF and ALT Strings must include the base before the event
(which must be reflected in the POS field), unless the event occurs at
position 1 on the contig in which case it must include the base after the
event; this padding base is not required (although it is permitted) for e.g.
complex substitutions or other events where all alleles have at least one
base represented in their Strings. If any of the ALT alleles is a symbolic
allele (an angle-bracketed ID String ``$<$ID$>$'') then the padding base is
required and POS denotes the coordinate of the base preceding the
polymorphism. Tools processing VCF files are not required to preserve case in
the allele Strings. (String, Required).
If the reference sequence contains IUPAC ambiguity codes not
allowed by this specification (such as R = A/G), the ambiguous reference base
must be reduced to a concrete base by using the one that is first alphabetically
(thus R as a reference base is converted to A in VCF.)
\item ALT - alternate base(s): Comma separated list of alternate non-reference alleles. These alleles do not have to be called in any of the samples. Options are base Strings made up of the bases A,C,G,T,N,*, (case insensitive) or an angle-bracketed ID String (``$<$ID$>$'') or a breakend replacement string as described in the section on breakends. The `*' allele is reserved to indicate that the allele is missing due to a an overlapping deletion. If there are no alternative alleles, then the missing value must be used. Tools processing VCF files are not required to preserve case in the allele String, except for IDs, which are case sensitive. (String; no whitespace, commas, or angle-brackets are permitted in the ID String itself)
\item QUAL - quality: Phred-scaled quality score for the assertion made in ALT. i.e. $-10log_{10}$ prob(call in ALT is wrong). If ALT is `.' (no variant) then this is $-10log_{10}$ prob(variant), and if ALT is not `.' this is $-10log_{10}$ prob(no variant). If unknown, the missing value must be specified. (Float)
\item FILTER - filter status: PASS if this position has passed all filters, i.e. a call is made at this position. Otherwise, if the site has not passed all filters, a semicolon-separated list of codes for filters that fail. e.g. ``q10;s50'' might indicate that at this site the quality is below 10 and the number of samples with data is below 50\% of the total number of samples. `0' is reserved and must not be used as a filter String. If filters have not been applied, then this field must be set to the missing value. (String, no white-space or semi-colons permitted, duplicate values not allowed.)
\item INFO - additional information: (String, no semi-colons or equals-signs permitted; commas are permitted only as delimiters for lists of
values; characters with special meaning can be encoded using the percent encoding, see Section~\ref{character-encoding}; space characters are allowed)
INFO fields are encoded as a semicolon-separated series of short keys with optional values in the format: $<$key$>$=$<$data$>$[,data].
INFO keys must match the regular expression \texttt{\^{}[A-Za-z\_][0-9A-Za-z\_.]*\$}, duplicate fields are not allowed. Arbitrary keys are permitted, although the sub-fields listed in Table~\ref{table:reserved-info} are reserved (albeit optional).
\begin{table}[htbp]
\centering
\begin{tabularx}{\textwidth}{ | l | l | l | X | }
Field & Number & Type & Description \\ \hline
AA & 1 & String & Ancestral allele \\
AC & A & Integer & Allele count in genotypes, for each ALT allele, in the same order as listed \\
AD & R & Integer & Total read depth for each allele \\
ADF & R & Integer & Read depth for each allele on the forward strand \\
ADR & R & Integer & Read depth for each allele on the reverse strand \\
AF & A & Float & Allele frequency for each ALT allele in the same order as listed (estimated from primary data, not called genotypes) \\
AN & 1 & Integer & Total number of alleles in called genotypes \\
BQ & 1 & Float & RMS base quality \\
CIGAR & A & String & Cigar string describing how to align an alternate allele to the reference allele \\
DB & 0 & Flag & dbSNP membership \\
DP & 1 & Integer & Combined depth across samples \\
END & 1 & Integer & End position (for use with symbolic alleles) \\
H2 & 0 & Flag & HapMap2 membership \\
H3 & 0 & Flag & HapMap3 membership \\
MQ & 1 & . & RMS mapping quality \\
MQ0 & 1 & Integer & Number of MAPQ == 0 reads \\
NS & 1 & Integer & Number of samples with data \\
SB & . & . & Strand bias \\
SOMATIC & 0 & Flag & Somatic mutation (for cancer genomics) \\
VALIDATED & 0 & Flag & Validated by follow-up experiment \\
1000G & 0 & Flag & 1000 Genomes membership \\
\end{tabularx}
\caption{Reserved INFO fields}
\label{table:reserved-info}
\end{table}
The exact format of each INFO sub-field should be specified in the meta-information (as described above).
Example for an INFO field: DP=154;MQ=52;H2. Keys without corresponding values may be used to indicate group membership (e.g. H2 indicates the SNP is found in HapMap 2). See Section~\ref{sv-info-keys} for additional reserved INFO sub-fields used to encode structural variants.
\end{enumerate}
\subsubsection{Genotype fields}
If genotype information is present, then the same types of data must be present
for all samples. First a FORMAT field is given specifying the data types and
order (colon-separated FORMAT ids matching the regular expression \texttt{\^{}[A-Za-z\_][0-9A-Za-z\_.]*\$}, duplicate fields are not allowed). This is followed by one data block per
sample, with the colon-separated data corresponding to the types
specified in the format. The first sub-field must always be the genotype (GT)
if it is present. There are no required sub-fields.
As with the INFO field, there are several common, reserved keywords that are standards across the community:
\begin{itemize}
\renewcommand{\labelitemii}{$\circ$}
\item AD, ADF, ADR: per-sample read depths for each allele; total (AD), on the forward (ADF) and the reverse (ADR) strand (Integer, Number=R)
\item DP : read depth at this position for this sample (Integer)
\item EC : comma separated list of expected alternate allele counts for each alternate allele in the same order as listed in the ALT field (typically used in association analyses) (Integers)
\item FT : sample genotype filter indicating if this genotype was ``called'' (similar in concept to the FILTER field). Again, use PASS to indicate that all filters have been passed, a semi-colon separated list of codes for filters that fail, or `.' to indicate that filters have not been applied. These values should be described in the meta-information in the same way as FILTERs (String, no white-space or semi-colons permitted)
\item GQ : conditional genotype quality, encoded as a phred quality $-10log_{10}$ p(genotype call is wrong, conditioned on the site's being variant) (Integer)
\item GP : genotype posterior probabilities in the range 0 to 1 using the same ordering as the GL field; one use can be to store imputed genotype probabilities (Float)
\item GT : genotype, encoded as allele values separated by either of $/$ or $\mid$. The allele values are 0 for the reference allele (what is in the REF field), 1 for the first allele listed in ALT, 2 for the second allele list in ALT and so on. For diploid calls examples could be $0/1$, $1\mid0$, or $1/2$, etc. Haploid calls, e.g. on Y, male non-pseudoautosomal X, or mitochondrion, are indicated by having only one allele value. A triploid call might look like $0/0/1$. If a call cannot be made for a sample at a given locus, `.' must be specified for each missing allele in the GT field (for example `$./.$' for a diploid genotype and `.' for haploid genotype). The meanings of the separators are as follows (see the PS field below for more details on incorporating phasing information into the genotypes):
\begin{itemize}
\item $/$ : genotype unphased
\item $\mid$ : genotype phased
\end{itemize}
\item GL : genotype likelihoods comprised of comma separated floating point
$log_{10}$-scaled likelihoods for all possible genotypes given the set of
alleles defined in the REF and ALT fields. In presence of the GT field the
same ploidy is expected; without GT field, diploidy is assumed.
\textsc{Genotype Ordering.} In general case of ploidy P and N alternate alleles (0 is the REF and 1..N
the alternate alleles), the ordering of genotypes for the likelihoods can
be expressed by the following pseudocode with as many nested loops as ploidy:\footnote{Note that we use inclusive \texttt{for} loop boundaries.}
\begingroup
\small
\begin{lstlisting}
for $a_P = 0\ldots N$
for $a_{P-1} = 0\ldots a_P$
$\ldots$
for $a_1 = 0\ldots a_{2}$
println $a_1 a_2 \ldots a_P$
\end{lstlisting}
\endgroup
Alternatively, the same can be achieved recursively with the following pseudocode:
\begingroup
\small
\begin{lstlisting}
Ordering($P$, $N$, suffix=""):
for $a$ in $0\ldots N$
if ($P == 1$) println str($a$) + suffix
if ($P > 1$) Ordering($P$-1, $a$, str($a$) + suffix)
\end{lstlisting}
\endgroup
Conversely, the index of the value corresponding to the genotype $k_1\le k_2\le\ldots\le k_P$ is
\begingroup
\small
\begin{lstlisting}
Index($k_1/k_2/\ldots/k_P$) = $\sum_{m=1}^{P} {k_m + m - 1 \choose m}$
\end{lstlisting}
\endgroup
Examples:
\begin{itemize}
\item for $P$=2 and $N$=1, the ordering is 00,01,11
\item for $P$=2 and $N$=2, the ordering is 00,01,11,02,12,22
\item for $P$=3 and $N$=2, the ordering is 000, 001, 011, 111, 002, 012, 112, 022, 122, 222
\item for $P$=1, the index of the genotype $a$ is $a$
\item for $P$=2, the index of the genotype "$a/b$", where $a\le b$, is $b (b+1)/2 + a$
\item for $P$=2 and arbitrary $N$, the ordering can be easily derived from a triangular matrix
\newline
\hbox{\hskip5em\footnotesize
\begin{tabular}{l|llll}
$b\setminus a$ & 0 & 1 & 2 & 3 \\ \hline \\[-0.5em]
0 & 0 & & & \\
1 & 1 & 2 & & \\
2 & 3 & 4 & 5 & \\
3 & 6 & 7 & 8 & 9
\end{tabular}
}
\end{itemize}
\item HQ : haplotype qualities, two comma separated phred qualities (Integers)
\item MQ : RMS mapping quality, similar to the version in the INFO field. (Integer)
\item PL : the phred-scaled genotype likelihoods rounded to the closest integer (and otherwise defined precisely as the GL field) (Integers)
\item PQ : phasing quality, the phred-scaled probability that alleles are ordered incorrectly in a heterozygote (against all other members in the phase set). We note that we have not yet included the specific measure for precisely defining ``phasing quality''; our intention for now is simply to reserve the PQ tag for future use as a measure of phasing quality. (Integer)
\item PS : phase set. A phase set is defined as a set of phased genotypes to which this genotype belongs. Phased genotypes for an individual that are on the same chromosome and have the same PS value are in the same phased set. A phase set specifies multi-marker haplotypes for the phased genotypes in the set. All phased genotypes that do not contain a PS subfield are assumed to belong to the same phased set. If the genotype in the GT field is unphased, the corresponding PS field is ignored. The recommended convention is to use the position of the first variant in the set as the PS identifier (although this is not required). (Non-negative 32-bit Integer)
\item $\ldots$ see Section~\ref{sv-format-keys} for a list of genotype keys reserved for structural variants.
\end{itemize}
If any of the fields is missing, it is replaced with the missing value. For example if the FORMAT is GT:GQ:DP:HQ then $0\mid0:.:23:23,34$ indicates that GQ is missing. Trailing fields can be dropped (with the exception of the GT field, which should always be present if specified in the FORMAT field).
See below for additional genotype fields used to encode structural variants. Additional Genotype fields can be defined in the meta-information. However, software support for such fields is not guaranteed.
\section{Understanding the VCF format and the haplotype representation}
VCF records use a single general system for representing genetic variation data composed of:
\begin{itemize}
\item Allele: representing single genetic haplotypes (A, T, ATC).
\item Genotype: an assignment of alleles for each chromosome of a single named sample at a particular locus.
\item VCF record: a record holding all segregating alleles at a locus (as well as genotypes, if appropriate, for multiple individuals containing alleles at that locus).
\end{itemize}
VCF records use a simple haplotype representation for REF and ALT alleles to describe variant haplotypes at a locus. ALT haplotypes are constructed from the REF haplotype by taking the REF allele bases at the POS in the reference genotype and replacing them with the ALT bases. In essence, the VCF record specifies a-REF-t and the alternative haplotypes are a-ALT-t for each alternative allele.
\subsection{VCF tag naming conventions}
\begin{itemize}
\item The "L" suffix means "likelihood" as log-likelihood in the sampling
distribution, log10 Pr(Data$|$Model). Likelihoods are represented as log10
scale, thus they are negative numbers (e.g.~GL, CNL). The likelihood can be also
represented in some cases as phred-scale in a separate tag (e.g.~PL).
\item The "P" suffix means "probability" as linear-scale probability in the
posterior distribution, which is Pr(Model$|$Data). Examples are GP, CNP.
\item The "Q" suffix means "quality" as log-complementary-phred-scale posterior
probability, which is -10 * log10 Pr(Data$|$Model) where the model is the most
likely genotype that appears in the GT field. Examples are GQ, CNQ. The fixed
site-level QUAL field follows the same convention (represented as a
phred-scaled number).
\end{itemize}
\section{INFO keys used for structural variants}
\label{sv-info-keys}
The following INFO keys are reserved for encoding structural variants. In general, when these keys are used by imprecise variants, the values should be best estimates. When a key reflects a property of a single alt allele (e.g. SVLEN), then when there are multiple alt alleles there will be multiple values for the key corresponding to each alelle (e.g. SVLEN=-100,-110 for a deletion with two distinct alt alleles).
\footnotesize
\begin{verbatim}
##INFO=<ID=IMPRECISE,Number=0,Type=Flag,Description="Imprecise structural variation">
##INFO=<ID=NOVEL,Number=0,Type=Flag,Description="Indicates a novel structural variation">
##INFO=<ID=END,Number=1,Type=Integer,Description="End position of the variant described in this record">
\end{verbatim}
\normalsize
For precise variants, END is POS + length of REF allele - 1, and the for imprecise variants the corresponding best estimate.
\footnotesize
\begin{verbatim}
##INFO=<ID=SVTYPE,Number=1,Type=String,Description="Type of structural variant">
\end{verbatim}
\normalsize
This key can be derived from the REF/ALT fields but is useful for filtering. The reserved values must be used for the types listed below:
\begin{itemize}
\item DEL: Deletion relative to the reference
\item INS: Insertion of novel sequence relative to the reference
\item DUP: Region of elevated copy number relative to the reference
\item INV: Inversion of reference sequence
\item CNV: Copy number variable region (may be both deletion and duplication)
\end{itemize}
\footnotesize
\begin{verbatim}
##INFO=<ID=SVLEN,Number=.,Type=Integer,Description="Difference in length between REF and ALT alleles">
\end{verbatim}
\normalsize
One value for each ALT allele. Longer ALT alleles (e.g. insertions) have positive values, shorter ALT alleles (e.g. deletions) have negative values.
\footnotesize
\begin{verbatim}
##INFO=<ID=CIPOS,Number=2,Type=Integer,Description="Confidence interval around POS for imprecise variants">
##INFO=<ID=CIEND,Number=2,Type=Integer,Description="Confidence interval around END for imprecise variants">
##INFO=<ID=HOMLEN,Number=.,Type=Integer,Description="Length of base pair identical micro-homology at event breakpoints">
##INFO=<ID=HOMSEQ,Number=.,Type=String,Description="Sequence of base pair identical micro-homology at event breakpoints">
##INFO=<ID=BKPTID,Number=.,Type=String,Description="ID of the assembled alternate allele in the assembly file">
\end{verbatim}
\normalsize
For precise variants, the consensus sequence the alternate allele assembly is derivable from the REF and ALT fields. However, the alternate allele assembly file may contain additional information about the characteristics of the alt allele contigs.
\footnotesize
\begin{verbatim}
##INFO=<ID=MEINFO,Number=4,Type=String,Description="Mobile element info of the form NAME,START,END,POLARITY">
##INFO=<ID=METRANS,Number=4,Type=String,Description="Mobile element transduction info of the form CHR,START,END,POLARITY">
##INFO=<ID=DGVID,Number=1,Type=String,Description="ID of this element in Database of Genomic Variation">
##INFO=<ID=DBVARID,Number=1,Type=String,Description="ID of this element in DBVAR">
##INFO=<ID=DBRIPID,Number=1,Type=String,Description="ID of this element in DBRIP">
##INFO=<ID=MATEID,Number=.,Type=String,Description="ID of mate breakends">
##INFO=<ID=PARID,Number=1,Type=String,Description="ID of partner breakend">
##INFO=<ID=EVENT,Number=1,Type=String,Description="ID of event associated to breakend">
##INFO=<ID=CILEN,Number=2,Type=Integer,Description="Confidence interval around the inserted material between breakends">
##INFO=<ID=DP,Number=1,Type=Integer,Description="Read Depth of segment containing breakend">
##INFO=<ID=DPADJ,Number=.,Type=Integer,Description="Read Depth of adjacency">
##INFO=<ID=CN,Number=1,Type=Integer,Description="Copy number of segment containing breakend">
##INFO=<ID=CNADJ,Number=.,Type=Integer,Description="Copy number of adjacency">
##INFO=<ID=CICN,Number=2,Type=Integer,Description="Confidence interval around copy number for the segment">
##INFO=<ID=CICNADJ,Number=.,Type=Integer,Description="Confidence interval around copy number for the adjacency">
\end{verbatim}
\normalsize
\section{FORMAT keys used for structural variants}
\label{sv-format-keys}
\footnotesize
\begin{verbatim}
##FORMAT=<ID=CN,Number=1,Type=Integer,Description="Copy number genotype for imprecise events">
##FORMAT=<ID=CNQ,Number=1,Type=Float,Description="Copy number genotype quality for imprecise events">
##FORMAT=<ID=CNL,Number=G,Type=Float,Description="Copy number genotype likelihood for imprecise events">
##FORMAT=<ID=CNP,Number=G,Type=Float,Description="Copy number posterior probabilities">
##FORMAT=<ID=NQ,Number=1,Type=Integer,Description="Phred style probability score that the variant is novel">
##FORMAT=<ID=HAP,Number=1,Type=Integer,Description="Unique haplotype identifier">
##FORMAT=<ID=AHAP,Number=1,Type=Integer,Description="Unique identifier of ancestral haplotype">
\end{verbatim}
\normalsize
These keys are analogous to GT/GQ/GL/GP and are provided for genotyping
imprecise events by copy number (either because there is an unknown number of
alternate alleles or because the haplotypes cannot be determined). CN specifies
the integer copy number of the variant in this sample. CNQ is encoded as a
phred quality $-10log_{10}$ p(copy number genotype call is wrong). CNL
specifies a list of $log_{10}$ likelihoods for each potential copy number,
starting from zero. CNP is 0 to 1-scaled copy number posterior
probabilities (and otherwise defined precisely as the CNL field), intended to
store imputed genotype probabilities. When possible, GT/GQ/GL/GP should be used
instead of (or in addition to) these keys.
\section{Representing variation in VCF records}
\subsection{Creating VCF entries for SNPs and small indels}
\subsubsection{Example 1}
For example, suppose we are looking at a locus in the genome:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & a t C g a & C is the reference base \\ \hline
1 & a t G g a & C base is a G in some individuals \\ \hline
2 & a t \ - \ g a & C base is deleted w.r.t. the reference sequence\\ \hline
3 & a t CAg a & A base is inserted w.r.t. the reference sequence \\ \hline
\end{tabular}
\vspace{0.3cm}
Representing these as VCF records would be done as follows:
\begin{enumerate}
\item A SNP polymorphism of C/G at position 3 where C is the reference base and G is the alternate base becomes REF=C, ALT=G
\item A single base deletion of C at position 3 becomes REF=TC, ALT=T
\item A single base insertion of A after position 3 becomes REF=C, ALT=CA
\end{enumerate}
Note that the positions must be sorted in increasing order:
\vspace{0.5em}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $2$ & . & TC & T & . & PASS & DP=100 \\
$20$ & $3$ & . & C & G & . & PASS & DP=100 \\
$20$ & $3$ & . & C & CA & . & PASS & DP=100 \\
\end{tabular}
\subsubsection{Example 2}
Suppose I see a the following in a population of individuals and want to represent these three segregating alleles:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & a t C g a & C is the reference base \\ \hline
$1$ & a t G g a & C base is a G in some individuals \\ \hline
$2$ & a t \ - \ g a & C base is deleted w.r.t. the reference sequence \\ \hline
\end{tabular}
\vspace{0.3cm}
In this case there are three segregating alleles: $\{tC,tG,t\}$ with a corresponding VCF record:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $2$ & . & TC & TG,T & . & PASS & DP=100 \\
\end{tabular}
\subsubsection{Example 3}
Now suppose I have this more complex example:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & a t C g a & C is the reference base \\ \hline
$1$ & a t \ - \ g a & C base is is deleted w.r.t. the reference sequence \\ \hline
$2$ & a t \ - - \ a & C and G bases are deleted w.r.t. the reference sequence\\ \hline
$3$ & a t CAg a & A base is inserted w.r.t. the reference sequence \\ \hline
\end{tabular}
\vspace{0.3cm}
There are actually four segregating alleles: $\{tCg,tg,t,tCAg\}$ over bases 2-4. This complex set of allele is represented in VCF as:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $2$ & . & TCG & TG,T,TCAG & . & PASS & DP=100 \\
\end{tabular}
\vspace{0.3cm}
Note that in VCF records, the molecular equivalence explicitly listed above in the per-base alignment is discarded, so the actual placement of equivalent g isn't retained. For completeness, VCF records are dynamically typed, so whether a VCF record is a SNP, Indel, Mixed, or Reference site depends on the properties of the alleles in the record.
\subsection{Decoding VCF entries for SNPs and small indels}
\subsubsection{SNP VCF record}
Suppose I receive the following VCF record:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $3$ & . & C & T & . & PASS & DP=100 \\
\end{tabular}
\vspace{0.3cm}
This is a SNP since its only single base substitution and there are only two alleles so I have the two following segregating haplotypes:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & \verb|a t C g a| & C is the reference base \\ \hline
$1$ & \verb|a t T g a| & C base is a T in some individuals \\ \hline
\end{tabular}
\subsubsection{Insertion VCF record}
Suppose I receive the following VCF record:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $3$ & . & C & CTAG & . & PASS & DP=100 \\
\end{tabular}
\vspace{0.3cm}
This is a insertion since the reference base C is being replaced by C [the reference base] plus three insertion bases TAG. Again there are only two alleles so I have the two following segregating haplotypes:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & \verb|a t C - - - g a| & C is the reference base \\ \hline
$1$ & \verb|a t C T A G g a| & following the C base is an insertion of 3 bases \\ \hline
\end{tabular}
\subsubsection{Deletion VCF record}
Suppose I receive the following VCF record:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $2$ & . & TCG & T & . & PASS & DP=100 \\
\end{tabular}
\vspace{0.3cm}
This is a deletion of two reference bases since the reference allele TCG is being replaced by just the T [the reference base]. Again there are only two alleles so I have the two following segregating haplotypes:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & \verb|a T C G a| & T is the (first) reference base \\ \hline
$1$ & \verb|a T - - a| & following the T base is a deletion of 2 bases \\ \hline
\end{tabular}
\subsubsection{Mixed VCF record for a microsatellite}
Suppose I receive the following VCF record:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l}
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$20$ & $4$ & . & GCG & G,GCGCG & . & PASS & DP=100 \\
\end{tabular}
\vspace{0.3cm}
This is a mixed type record containing a 2 base insertion and a 2 base deletion. There are are three segregating alleles so I have the three following haplotypes:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & \verb|a t c G C G - - a| & G is the (first) reference base \\ \hline
$1$ & \verb|a t c G - - - - a| & following the G base is a deletion of 2 bases \\ \hline
$2$ & \verb|a t c G C G C G a| & following the G base is a insertion of 2 bases \\ \hline
\end{tabular}
\vspace{0.3cm}
Note that in all of these examples dashes have been added to make the haplotypes clearer but of course the equivalence among bases isn't provided by the VCF. Technically the following is an equivalent alignment:
\vspace{0.3cm}
\begin{tabular}{ | l | l | l | }
\hline
Example & Sequence & Alteration \\ \hline
Ref & \verb|a t c G - - C G a| & G is the (first) reference base \\ \hline
$1$ & \verb|a t c G - - - - a| & following the G base is a deletion of 2 bases \\ \hline
$2$ & \verb|a t c G C G C G a| & following the G base is a insertion of 2 bases \\ \hline
\end{tabular}
\subsection{Encoding Structural Variants}
The following page contains examples of structural variants encoded in VCF:
\pagebreak
\footnotesize
\begin{landscape}
\begin{verbatim}
VCF STRUCTURAL VARIANT EXAMPLE
##fileformat=VCFv4.1
##fileDate=20100501
##reference=1000GenomesPilot-NCBI36
##assembly=ftp://ftp-trace.ncbi.nih.gov/1000genomes/ftp/release/sv/breakpoint_assemblies.fasta
##INFO=<ID=BKPTID,Number=.,Type=String,Description="ID of the assembled alternate allele in the assembly file">
##INFO=<ID=CIEND,Number=2,Type=Integer,Description="Confidence interval around END for imprecise variants">
##INFO=<ID=CIPOS,Number=2,Type=Integer,Description="Confidence interval around POS for imprecise variants">
##INFO=<ID=END,Number=1,Type=Integer,Description="End position of the variant described in this record">
##INFO=<ID=HOMLEN,Number=.,Type=Integer,Description="Length of base pair identical micro-homology at event breakpoints">
##INFO=<ID=HOMSEQ,Number=.,Type=String,Description="Sequence of base pair identical micro-homology at event breakpoints">
##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">
##ALT=<ID=DEL,Description="Deletion">
##ALT=<ID=DEL:ME:ALU,Description="Deletion of ALU element">
##ALT=<ID=DEL:ME:L1,Description="Deletion of L1 element">
##ALT=<ID=DUP,Description="Duplication">
##ALT=<ID=DUP:TANDEM,Description="Tandem Duplication">
##ALT=<ID=INS,Description="Insertion of novel sequence">
##ALT=<ID=INS:ME:ALU,Description="Insertion of ALU element">
##ALT=<ID=INS:ME:L1,Description="Insertion of L1 element">
##ALT=<ID=INV,Description="Inversion">
##ALT=<ID=CNV,Description="Copy number variable region">
##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">
##FORMAT=<ID=GQ,Number=1,Type=Float,Description="Genotype quality">
##FORMAT=<ID=CN,Number=1,Type=Integer,Description="Copy number genotype for imprecise events">
##FORMAT=<ID=CNQ,Number=1,Type=Float,Description="Copy number genotype quality for imprecise events">
#CHROM POS ID REF ALT QUAL FILTER INFO FORMAT NA00001
1 2827694 rs2376870 CGTGGATGCGGGGAC C . PASS SVTYPE=DEL;END=2827708;HOMLEN=1;HOMSEQ=G;SVLEN=-14 GT:GQ 1/1:13.9
2 321682 . T <DEL> 6 PASS SVTYPE=DEL;END=321887;SVLEN=-205;CIPOS=-56,20;CIEND=-10,62 GT:GQ 0/1:12
2 14477084 . C <DEL:ME:ALU> 12 PASS SVTYPE=DEL;END=14477381;SVLEN=-297;CIPOS=-22,18;CIEND=-12,32 GT:GQ 0/1:12
3 9425916 . C <INS:ME:L1> 23 PASS SVTYPE=INS;END=9425916;SVLEN=6027;CIPOS=-16,22 GT:GQ 1/1:15
3 12665100 . A <DUP> 14 PASS SVTYPE=DUP;END=12686200;SVLEN=21100;CIPOS=-500,500;CIEND=-500,500 GT:GQ:CN:CNQ ./.:0:3:16.2
4 18665128 . T <DUP:TANDEM> 11 PASS SVTYPE=DUP;END=18665204;SVLEN=76;CIPOS=-10,10;CIEND=-10,10 GT:GQ:CN:CNQ ./.:0:5:8.3
\end{verbatim}
\end{landscape}
\pagebreak
\normalsize
The example shows in order:
\begin{enumerate}
\item A precise deletion with known breakpoint, a one base micro-homology, and a sample that is homozygous for the deletion.
\item An imprecise deletion of approximately 205 bp.
\item An imprecise deletion of an ALU element relative to the reference.
\item An imprecise insertion of an L1 element relative to the reference.
\item An imprecise duplication of approximately 21Kb. The sample genotype is copy number 3 (one extra copy of the duplicated sequence).
\item An imprecise tandem duplication of 76bp. The sample genotype is copy number 5 (but the two haplotypes are not known).
\end{enumerate}
\subsection{Specifying complex rearrangements with breakends}
An arbitrary rearrangement event can be summarized as a set of novel \textbf{adjacencies}. Each adjacency ties together $2$ \textbf{breakends}. The two breakends at either end of a novel adjacency are called \textbf{mates}.
There is one line of VCF (i.e. one record) for each of the two breakends in a novel adjacency. A breakend record is identified with the tag ``SVTYPE=BND'' in the INFO field. The REF field of a breakend record indicates a base or sequence s of bases beginning at position POS, as in all VCF records. The ALT field of a breakend record indicates a replacement for s. This ``breakend replacement'' has three parts:
\begin{enumerate}
\item The string t that replaces places s. The string t may be an extended version of s if some novel bases are inserted during the formation of the novel adjacency.
\item The position p of the mate breakend, indicated by a string of the form ``chr:pos''. This is the location of the first mapped base in the piece being joined at this novel adjacency.
\item The direction that the joined sequence continues in, starting from p. This is indicated by the orientation of square brackets surrounding p.
\end{enumerate}
These 3 elements are combined in 4 possible ways to create the ALT. In each of the 4 cases, the assertion is that s is replaced with t, and then some piece starting at position p is joined to t. The cases are:
\vspace{0.3cm}
\begin{tabular}{ l l l }
REF & ALT & Meaning \\
s & t$[$p$[$ & piece extending to the right of p is joined after t \\
s & t$]$p$]$ & reverse comp piece extending left of p is joined after t \\
s & $]$p$]$t & piece extending to the left of p is joined before t \\
s & $[$p$[$t & reverse comp piece extending right of p is joined before t \\
\end{tabular}
\vspace{0.3cm}
The example in Figure 1 shows a 3-break operation involving 6 breakends. It exemplifies all possible orientations of breakends in adjacencies. Notice how the ALT field expresses the orientation of the breakends.
\begin{figure}[ht]
\centering
\includegraphics[width=4in,height=2.96in]{img/all_orientations-400x296.png}
\caption{All possible orientations of breakends}
\end{figure}
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l }
\#CHROM &POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$2$ & $321681$ & bnd\_W & G & G$]17$:$198982]$ & $6$ & PASS & SVTYPE=BND \\
$2$ & $321682$ & bnd\_V & T & $]$13:123456$]$T & 6 & PASS & SVTYPE=BND \\
$13$ & $123456$ & bnd\_U & C & C$[$2:321682$[$ & 6 & PASS & SVTYPE=BND \\
$13$ & $123457$ & bnd\_X & A & $[$17:198983$[$A & 6 & PASS & SVTYPE=BND \\
$17$ & $198982$ & bnd\_Y & A & A$]$2:321681$]$ & 6 & PASS & SVTYPE=BND \\
$17$ & $198983$ & bnd\_Z & C & $[$13:123457$[$C & 6 & PASS & SVTYPE=BND \\
\end{tabular}
\subsubsection{Inserted Sequence}
Sometimes, as shown in Figure 2, some bases are inserted between the two breakends, this information is also carried in the ALT column:
\begin{figure}[h]
\centering
\includegraphics[width=4in,height=1.89in]{img/inserted_sequence-400x189.png}
\caption{Inserted sequence between breakends}
\end{figure}
\vspace{0.3cm}
\footnotesize
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$2$ & $321682$ & bnd\_V & T & $]13:123456]$AGTNNNNNCAT & $6$ & PASS & SVTYPE=BND;MATEID=bnd\_U \\
$13$ & $123456$ & bnd\_U & C & CAGTNNNNNCA$[2:321682[$ & $6$ & PASS & SVTYPE=BND;MATEID=bnd\_V \\
\end{tabular}
\normalsize
\vspace{0.3cm}
\subsubsection{Large Insertions}
If the insertion is too long to be conveniently stored in the ALT column, as in the 329 base insertion shown in Figure 3, it can be represented by a contig from the assembly file:
\begin{figure}[h]
\centering
\includegraphics[width=4in,height=2.47in]{img/inserted_contig-400x247.png}
\caption{Inserted contig}
\end{figure}
\vspace{0.3cm}
\small
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$13$ & $123456$ & bnd\_U & C & C$[<$ctg1$>:1[$ & $6$ & PASS & SVTYPE=BND \\
$13$ & $123457$ & bnd\_V & A & $]<$ctg$1>:329]$A & $6$ & PASS & SVTYPE=BND \\
\end{tabular}
\normalsize
\vspace{0.3cm}
\textbf{Note}: In the special case of the complete insertion of a sequence between two base pairs, it is recommended to use the shorthand notation described above:
\vspace{0.3cm}
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$13$ & $321682$ & INS0 & T & C$<$ctg$1>$ & $6$ & PASS & SVTYPE=INS \\
\end{tabular}
\vspace{0.3cm}
If only a portion of $<$ctg$1>$, say from position $7$ to position $214$, is inserted, the VCF would be:
\vspace{0.3cm}
\small
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$13$ & $123456$ & bnd\_U & C & C$[<$ctg1$>:7[$ & $6$ & PASS & SVTYPE=BND \\
$13$ & $123457$ & bnd\_V & A & $]<$ctg$1>:214]$A & $6$ & PASS & SVTYPE=BND \\
\end{tabular}
\normalsize
\vspace{0.3cm}
If $<$ctg$1>$ is circular and a segment from position 229 to position 45 is inserted, i.e. continuing from position 329 on to position 1, this is represented by adding a circular adjacency:
\vspace{0.3cm}
\small
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$13$ & $123456$ & bnd\_U & C & C$[<$ctg$1>:229[$ & 6 & PASS & SVTYPE=BND \\
$13$ & $123457$ & bnd\_V & A & $]<$ctg$1>:45]$A & 6 & PASS & SVTYPE=BND \\
$<$ctg$1>$ & 1 & bnd\_X & A & $]<$ctg$1>:329]$A & 6 & PASS & SVTYPE=BND \\
$<$ctg$1>$ & 329 & bnd\_Y & T & T$[<$ctg$1>:1[$ & 6 & PASS & SVTYPE=BND \\
\end{tabular}
\normalsize
\subsubsection{Multiple mates}
If a breakend has multiple mates such as in Figure 4 (either because of breakend reuse or of uncertainty in the measurement), these alternate adjacencies are treated as alternate alleles:
\begin{figure}[h]
\centering
\includegraphics[width=4in,height=2.80in]{img/multiple_mates-400x280.png}
\caption{Breakend with multiple mates}
\end{figure}
\footnotesize
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
$2$ & $321682$ & bnd\_V & T & $]13:123456]$T & 6 & PASS & SVTYPE=BND;MATEID=bnd\_U \\
$13$ & $123456$ & bnd\_U & C & C$[2:321682[$,C$[17:198983[$ & 6 & PASS & SVTYPE=BND;MATEID=bnd\_V,bnd\_Z \\
$17$ & $198983$ & bnd\_Z & A & $]13:123456]$A & 6 & PASS & SVTYPE=BND;MATEID=bnd\_U \\
\end{tabular}
\normalsize
\subsubsection{Explicit partners}
Two breakends which are connected in the reference genome but disconnected in the variants are called partners. Each breakend only has one partner, typically one basepair left or right. However, it is not uncommon to observe loss of a few basepairs during the rearrangement. A breakend's partner may be explicitly named as in Figure 5:
\begin{figure}[ht]
\centering
\includegraphics[width=4in,height=2.11in]{img/erosion-400x211.png}
\caption{Partner breakends}
\end{figure}
\small
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
2 & 321681 & bnd\_W & G & G$[13:123460[$ & 6 & PASS & PARID=bnd\_V;MATEID=bnd\_X \\
2 & 321682 & bnd\_V & T & $]13:123456]$T & 6 & PASS & PARID=bnd\_W;MATEID=bnd\_U \\
13 & 123456 & bnd\_U & C & C$[2:321682[$ & 6 & PASS & PARID=bnd\_X;MATEID=bnd\_V \\
13 & 123460 & bnd\_X & A & $]2:321681]$A & 6 & PASS & PARID=bnd\_U;MATEID=bnd\_W \\
\end{tabular}
\normalsize
\subsubsection{Telomeres}
For a rearrangement involving the telomere end of a reference chromosome, we define a virtual telomeric breakend that serves as a breakend partner for the breakend at the telomere. That way every breakend has a partner. If the chromosome extends from position 1 to N, then the virtual telomeric breakends are at positions 0 and N+1. For example, to describe the reciprocal translocation of the entire chromosome 1 into chromosome 13, as illustrated in Figure 6:
\begin{figure}[h]
\centering
\includegraphics[width=4in,height=2.51in]{img/telomere-400x251.png}
\caption{Telomeres}
\end{figure}
the records would look like:
\small
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
1 & 0 & bnd\_X & N & $.[13:123457[$ & 6 & PASS & SVTYPE=BND;MATEID=bnd\_V \\
1 & 1 & bnd\_Y & T & $]13:123456]$T & 6 & PASS & SVTYPE=BND;MATEID=bnd\_U \\
13 & 123456 & bnd\_U & C & C$[1:1[$ & 6 & PASS & SVTYPE=BND;MATEID=bnd\_Y \\
13 & 123457 & bnd\_V & A & $]1:0]$A & 6 & PASS & SVTYPE=BND;MATEID=bnd\_X \\
\end{tabular}
\normalsize
\subsubsection{Event modifiers}
As mentioned previously, a single rearrangement event can be described as a set of novel adjacencies. For example, a reciprocal rearrangement such as in Figure 7:
\begin{figure}[h]
\centering
\includegraphics[width=4in,height=1.92in]{img/reciprocal_rearrangement-400x192.png}
\caption{Rearrangements}
\end{figure}
would be described as:
\vspace{0.3cm}
\footnotesize
\begin{tabular}{ l l l l l l l l }
\#CHROM & POS & ID & REF & ALT & QUAL & FILTER & INFO \\
2 & 321681 & bnd\_W & G & G$[13:123457[$ & 6 & PASS & SVTYPE=BND;MATEID=bnd\_X;EVENT=RR0 \\
2 & 321682 & bnd\_V & T & $]13:123456]$T & 6 & PASS & SVTYPE=BND;MATEID=bnd\_U;EVENT=RR0 \\
13 & 123456 & bnd\_U & C & C$[2:321682[$ & 6 & PASS & SVTYPE=BND;MATEID=bnd\_V;EVENT=RR0 \\
13 & 123457 & bnd\_X & A & $]2:321681]$A & 6 & PASS & SVTYPE=BND;MATEID=bnd\_W;EVENT=RR0 \\
\end{tabular}
\normalsize