-.TH samtools 1 "27 October 2010" "samtools-0.1.9" "Bioinformatics tools"
+.TH samtools 1 "05 July 2011" "samtools-0.1.17" "Bioinformatics tools"
.SH NAME
.PP
samtools - Utilities for the Sequence Alignment/Map (SAM) format
+
+bcftools - Utilities for the Binary Call Format (BCF) and VCF
.SH SYNOPSIS
.PP
samtools view -bt ref_list.txt -o aln.bam aln.sam.gz
.PP
samtools pileup -vcf ref.fasta aln.sorted.bam
.PP
-samtools mpileup -C50 -agf ref.fasta -r chr3:1,000-2,000 in1.bam in2.bam
+samtools mpileup -C50 -gf ref.fasta -r chr3:1,000-2,000 in1.bam in2.bam
.PP
samtools tview aln.sorted.bam ref.fasta
+.PP
+bcftools index in.bcf
+.PP
+bcftools view in.bcf chr2:100-200 > out.vcf
+.PP
+bcftools view -vc in.bcf > out.vcf 2> out.afs
.SH DESCRIPTION
.PP
will download the index upon absence. Samtools does not retrieve the
entire alignment file unless it is asked to do so.
-.SH COMMANDS AND OPTIONS
+.SH SAMTOOLS COMMANDS AND OPTIONS
.TP 10
.B view
-samtools view [-bhuHS] [-t in.refList] [-o output] [-f reqFlag] [-F
+samtools view [-bchuHS] [-t in.refList] [-o output] [-f reqFlag] [-F
skipFlag] [-q minMapQ] [-l library] [-r readGroup] [-R rgFile] <in.bam>|<in.sam> [region1 [...]]
Extract/print all or sub alignments in SAM or BAM format. If no region
.B -b
Output in the BAM format.
.TP
-.BI -f " INT"
+.BI -f \ INT
Only output alignments with all bits in INT present in the FLAG
field. INT can be in hex in the format of /^0x[0-9A-F]+/ [0]
.TP
-.BI -F " INT"
+.BI -F \ INT
Skip alignments with bits present in INT [0]
.TP
.B -h
.B -H
Output the header only.
.TP
-.BI -l " STR"
+.BI -l \ STR
Only output reads in library STR [null]
.TP
-.BI -o " FILE"
+.BI -o \ FILE
Output file [stdout]
.TP
-.BI -q " INT"
+.BI -q \ INT
Skip alignments with MAPQ smaller than INT [0]
.TP
-.BI -r " STR"
+.BI -r \ STR
Only output reads in read group STR [null]
.TP
-.BI -R " FILE"
+.BI -R \ FILE
Output reads in read groups listed in
.I FILE
[null]
.B `-t'
option is required.
.TP
-.BI -t " FILE"
+.B -c
+Instead of printing the alignments, only count them and print the
+total number. All filter options, such as
+.B `-f',
+.B `-F'
+and
+.B `-q'
+, are taken into account.
+.TP
+.BI -t \ FILE
This file is TAB-delimited. Each line must contain the reference name
and the length of the reference, one line for each distinct reference;
additional fields are ignored. This file also defines the order of the
viewing the same reference sequence.
.TP
-.B pileup
-samtools pileup [-2sSBicv] [-f in.ref.fasta] [-t in.ref_list] [-l
-in.site_list] [-C capMapQ] [-M maxMapQ] [-T theta] [-N nHap] [-r
-pairDiffRate] [-m mask] [-d maxIndelDepth] [-G indelPrior]
-<in.bam>|<in.sam>
+.B mpileup
+.B samtools mpileup
+.RB [ \-EBug ]
+.RB [ \-C
+.IR capQcoef ]
+.RB [ \-r
+.IR reg ]
+.RB [ \-f
+.IR in.fa ]
+.RB [ \-l
+.IR list ]
+.RB [ \-M
+.IR capMapQ ]
+.RB [ \-Q
+.IR minBaseQ ]
+.RB [ \-q
+.IR minMapQ ]
+.I in.bam
+.RI [ in2.bam
+.RI [ ... ]]
+
+Generate BCF or pileup for one or multiple BAM files. Alignment records
+are grouped by sample identifiers in @RG header lines. If sample
+identifiers are absent, each input file is regarded as one sample.
-Print the alignment in the pileup format. In the pileup format, each
+In the pileup format (without
+.BR -u or -g ),
+each
line represents a genomic position, consisting of chromosome name,
coordinate, reference base, read bases, read qualities and alignment
mapping qualities. Information on match, mismatch, indel, strand,
following `^' minus 33 gives the mapping quality. A symbol `$' marks the
end of a read segment.
-If option
-.B -c
-is applied, the consensus base, Phred-scaled consensus quality, SNP
-quality (i.e. the Phred-scaled probability of the consensus being
-identical to the reference) and root mean square (RMS) mapping quality
-of the reads covering the site will be inserted between the `reference
-base' and the `read bases' columns. An indel occupies an additional
-line. Each indel line consists of chromosome name, coordinate, a star,
-the genotype, consensus quality, SNP quality, RMS mapping quality, #
-covering reads, the first alllele, the second allele, # reads supporting
-the first allele, # reads supporting the second allele and # reads
-containing indels different from the top two alleles.
-
-The position of indels is offset by -1.
-
-.B OPTIONS:
+.B Input Options:
.RS
.TP 10
-.B -B
-Disable the BAQ computation. See the
-.B mpileup
-command for details.
+.B -6
+Assume the quality is in the Illumina 1.3+ encoding.
+.B -A
+Do not skip anomalous read pairs in variant calling.
.TP
-.B -c
-Call the consensus sequence using SOAPsnp consensus model. Options
-.BR -T ", " -N ", " -I " and " -r
-are only effective when
-.BR -c " or " -g
-is in use.
-.TP
-.BI -C " INT"
-Coefficient for downgrading the mapping quality of poorly mapped
-reads. See the
-.B mpileup
-command for details. [0]
-.TP
-.BI -d " INT"
-Use the first
-.I NUM
-reads in the pileup for indel calling for speed up. Zero for unlimited. [1024]
-.TP
-.BI -f " FILE"
-The reference sequence in the FASTA format. Index file
-.I FILE.fai
-will be created if
-absent.
-.TP
-.B -g
-Generate genotype likelihood in the binary GLFv3 format. This option
-suppresses -c, -i and -s. This option is deprecated by the
-.B mpileup
-command.
-.TP
-.B -i
-Only output pileup lines containing indels.
-.TP
-.BI -I " INT"
-Phred probability of an indel in sequencing/prep. [40]
-.TP
-.BI -l " FILE"
-List of sites at which pileup is output. This file is space
-delimited. The first two columns are required to be chromosome and
-1-based coordinate. Additional columns are ignored. It is
-recommended to use option
-.TP
-.BI -m " INT"
-Filter reads with flag containing bits in
-.I INT
-[1796]
-.TP
-.BI -M " INT"
-Cap mapping quality at INT [60]
-.TP
-.BI -N " INT"
-Number of haplotypes in the sample (>=2) [2]
-.TP
-.BI -r " FLOAT"
-Expected fraction of differences between a pair of haplotypes [0.001]
-.TP
-.B -s
-Print the mapping quality as the last column. This option makes the
-output easier to parse, although this format is not space efficient.
-.TP
-.B -S
-The input file is in SAM.
-.TP
-.BI -t " FILE"
-List of reference names ane sequence lengths, in the format described
-for the
-.B import
-command. If this option is present, samtools assumes the input
-.I <in.alignment>
-is in SAM format; otherwise it assumes in BAM format.
-.B -s
-together with
-.B -l
-as in the default format we may not know the mapping quality.
-.TP
-.BI -T " FLOAT"
-The theta parameter (error dependency coefficient) in the maq consensus
-calling model [0.85]
-.RE
-
-.TP
-.B mpileup
-samtools mpileup [-Bug] [-C capQcoef] [-r reg] [-f in.fa] [-l list] [-M capMapQ] [-Q minBaseQ] [-q minMapQ] in.bam [in2.bam [...]]
-
-Generate BCF or pileup for one or multiple BAM files. Alignment records
-are grouped by sample identifiers in @RG header lines. If sample
-identifiers are absent, each input file is regarded as one sample.
-
-.B OPTIONS:
-.RS
-.TP 8
.B -B
Disable probabilistic realignment for the computation of base alignment
quality (BAQ). BAQ is the Phred-scaled probability of a read base being
misaligned. Applying this option greatly helps to reduce false SNPs
caused by misalignments.
.TP
-.BI -C " INT"
+.BI -b \ FILE
+List of input BAM files, one file per line [null]
+.TP
+.BI -C \ INT
Coefficient for downgrading mapping quality for reads containing
excessive mismatches. Given a read with a phred-scaled probability q of
being generated from the mapped position, the new mapping quality is
about sqrt((INT-q)/INT)*INT. A zero value disables this
-functionality; if enabled, the recommended value is 50. [0]
+functionality; if enabled, the recommended value for BWA is 50. [0]
.TP
-.BI -f " FILE"
-The reference file [null]
+.BI -d \ INT
+At a position, read maximally
+.I INT
+reads per input BAM. [250]
.TP
-.B -g
-Compute genotype likelihoods and output them in the binary call format (BCF).
+.B -E
+Extended BAQ computation. This option helps sensitivity especially for MNPs, but may hurt
+specificity a little bit.
.TP
-.B -u
-Similar to
-.B -g
-except that the output is uncompressed BCF, which is preferred for pipeing.
+.BI -f \ FILE
+The
+.BR faidx -indexed
+reference file in the FASTA format. The file can be optionally compressed by
+.BR razip .
+[null]
.TP
-.BI -l " FILE"
-File containing a list of sites where pileup or BCF is outputted [null]
+.BI -l \ FILE
+BED or position list file containing a list of regions or sites where pileup or BCF should be generated [null]
.TP
-.BI -q " INT"
+.BI -q \ INT
Minimum mapping quality for an alignment to be used [0]
.TP
-.BI -Q " INT"
+.BI -Q \ INT
Minimum base quality for a base to be considered [13]
.TP
-.BI -r " STR"
+.BI -r \ STR
Only generate pileup in region
.I STR
[all sites]
+.TP
+.B Output Options:
+
+.TP
+.B -D
+Output per-sample read depth
+.TP
+.B -g
+Compute genotype likelihoods and output them in the binary call format (BCF).
+.TP
+.B -S
+Output per-sample Phred-scaled strand bias P-value
+.TP
+.B -u
+Similar to
+.B -g
+except that the output is uncompressed BCF, which is preferred for piping.
+
+.TP
+.B Options for Genotype Likelihood Computation (for -g or -u):
+
+.TP
+.BI -e \ INT
+Phred-scaled gap extension sequencing error probability. Reducing
+.I INT
+leads to longer indels. [20]
+.TP
+.BI -h \ INT
+Coefficient for modeling homopolymer errors. Given an
+.IR l -long
+homopolymer
+run, the sequencing error of an indel of size
+.I s
+is modeled as
+.IR INT * s / l .
+[100]
+.TP
+.B -I
+Do not perform INDEL calling
+.TP
+.BI -L \ INT
+Skip INDEL calling if the average per-sample depth is above
+.IR INT .
+[250]
+.TP
+.BI -o \ INT
+Phred-scaled gap open sequencing error probability. Reducing
+.I INT
+leads to more indel calls. [40]
+.TP
+.BI -P \ STR
+Comma dilimited list of platforms (determined by
+.BR @RG-PL )
+from which indel candidates are obtained. It is recommended to collect
+indel candidates from sequencing technologies that have low indel error
+rate such as ILLUMINA. [all]
.RE
.TP
This command is much faster than replacing the header with a
BAM->SAM->BAM conversion.
+.TP
+.B cat
+samtools cat [-h header.sam] [-o out.bam] <in1.bam> <in2.bam> [ ... ]
+
+Concatenate BAMs. The sequence dictionary of each input BAM must be identical,
+although this command does not check this. This command uses a similar trick
+to
+.B reheader
+which enables fast BAM concatenation.
+
.TP
.B sort
samtools sort [-no] [-m maxMem] <in.bam> <out.prefix>
.B -n
Sort by read names rather than by chromosomal coordinates
.TP
-.B -m INT
+.BI -m \ INT
Approximately the maximum required memory. [500000000]
.RE
.TP
.B merge
-samtools merge [-nur] [-h inh.sam] [-R reg] <out.bam> <in1.bam> <in2.bam> [...]
+samtools merge [-nur1f] [-h inh.sam] [-R reg] <out.bam> <in1.bam> <in2.bam> [...]
Merge multiple sorted alignments.
The header reference lists of all the input BAM files, and the @SQ headers of
.B OPTIONS:
.RS
.TP 8
-.BI -h " FILE"
+.B -1
+Use zlib compression level 1 to comrpess the output
+.TP
+.B -f
+Force to overwrite the output file if present.
+.TP 8
+.BI -h \ FILE
Use the lines of
.I FILE
as `@' headers to be copied to
is actually in SAM format, though any alignment records it may contain
are ignored.)
.TP
-.BI -R " STR"
+.B -n
+The input alignments are sorted by read names rather than by chromosomal
+coordinates
+.TP
+.BI -R \ STR
Merge files in the specified region indicated by
.I STR
+[null]
.TP
.B -r
Attach an RG tag to each alignment. The tag value is inferred from file names.
.TP
-.B -n
-The input alignments are sorted by read names rather than by chromosomal
-coordinates
-.TP
.B -u
Uncompressed BAM output
.RE
.TP
.B calmd
-samtools calmd [-eubSr] [-C capQcoef] <aln.bam> <ref.fasta>
+samtools calmd [-
EeubSr] [-C capQcoef] <aln.bam> <ref.fasta>
Generate the MD tag. If the MD tag is already present, this command will
give a warning if the MD tag generated is different from the existing
.B OPTIONS:
.RS
.TP 8
+.B -A
+When used jointly with
+.B -r
+this option overwrites the original base quality.
+.TP 8
.B -e
Convert a the read base to = if it is identical to the aligned reference
base. Indel caller does not support the = bases at the moment.
.B -S
The input is SAM with header lines
.TP
-.BI -C " INT"
+.BI -C \ INT
Coefficient to cap mapping quality of poorly mapped reads. See the
.B pileup
command for details. [0]
.TP
.B -r
-Perform probabilistic realignment to compute BAQ, which will be used to
-cap base quality.
+Compute the BQ tag (without -A) or cap base quality by BAQ (with -A).
+.TP
+.B -E
+Extended BAQ calculation. This option trades specificity for sensitivity, though the
+effect is minor.
+.RE
+
+.TP
+.B targetcut
+samtools targetcut [-Q minBaseQ] [-i inPenalty] [-0 em0] [-1 em1] [-2 em2] [-f ref] <in.bam>
+
+This command identifies target regions by examining the continuity of read depth, computes
+haploid consensus sequences of targets and outputs a SAM with each sequence corresponding
+to a target. When option
+.B -f
+is in use, BAQ will be applied. This command is
+.B only
+designed for cutting fosmid clones from fosmid pool sequencing [Ref. Kitzman et al. (2010)].
.RE
+.TP
+.B phase
+samtools phase [-AF] [-k len] [-b prefix] [-q minLOD] [-Q minBaseQ] <in.bam>
+
+Call and phase heterozygous SNPs.
+.B OPTIONS:
+.RS
+.TP 8
+.B -A
+Drop reads with ambiguous phase.
+.TP 8
+.BI -b \ STR
+Prefix of BAM output. When this option is in use, phase-0 reads will be saved in file
+.BR STR .0.bam
+and phase-1 reads in
+.BR STR .1.bam.
+Phase unknown reads will be randomly allocated to one of the two files. Chimeric reads
+with switch errors will be saved in
+.BR STR .chimeric.bam.
+[null]
+.TP
+.B -F
+Do not attempt to fix chimeric reads.
+.TP
+.BI -k \ INT
+Maximum length for local phasing. [13]
+.TP
+.BI -q \ INT
+Minimum Phred-scaled LOD to call a heterozygote. [40]
+.TP
+.BI -Q \ INT
+Minimum base quality to be used in het calling. [13]
+.RE
+
+.SH BCFTOOLS COMMANDS AND OPTIONS
+
+.TP 10
+.B view
+.B bcftools view
+.RB [ \-AbFGNQSucgv ]
+.RB [ \-D
+.IR seqDict ]
+.RB [ \-l
+.IR listLoci ]
+.RB [ \-s
+.IR listSample ]
+.RB [ \-i
+.IR gapSNPratio ]
+.RB [ \-t
+.IR mutRate ]
+.RB [ \-p
+.IR varThres ]
+.RB [ \-P
+.IR prior ]
+.RB [ \-1
+.IR nGroup1 ]
+.RB [ \-d
+.IR minFrac ]
+.RB [ \-U
+.IR nPerm ]
+.RB [ \-X
+.IR permThres ]
+.RB [ \-T
+.IR trioType ]
+.I in.bcf
+.RI [ region ]
+
+Convert between BCF and VCF, call variant candidates and estimate allele
+frequencies.
+
+.RS
+.TP
+.B Input/Output Options:
+.TP 10
+.B -A
+Retain all possible alternate alleles at variant sites. By default, the view
+command discards unlikely alleles.
+.TP 10
+.B -b
+Output in the BCF format. The default is VCF.
+.TP
+.BI -D \ FILE
+Sequence dictionary (list of chromosome names) for VCF->BCF conversion [null]
+.TP
+.B -F
+Indicate PL is generated by r921 or before (ordering is different).
+.TP
+.B -G
+Suppress all individual genotype information.
+.TP
+.BI -l \ FILE
+List of sites at which information are outputted [all sites]
+.TP
+.B -N
+Skip sites where the REF field is not A/C/G/T
+.TP
+.B -Q
+Output the QCALL likelihood format
+.TP
+.BI -s \ FILE
+List of samples to use. The first column in the input gives the sample names
+and the second gives the ploidy, which can only be 1 or 2. When the 2nd column
+is absent, the sample ploidy is assumed to be 2. In the output, the ordering of
+samples will be identical to the one in
+.IR FILE .
+[null]
+.TP
+.B -S
+The input is VCF instead of BCF.
+.TP
+.B -u
+Uncompressed BCF output (force -b).
+.TP
+.B Consensus/Variant Calling Options:
+.TP 10
+.B -c
+Call variants using Bayesian inference. This option automatically invokes option
+.BR -e .
+.TP
+.BI -d \ FLOAT
+When
+.B -v
+is in use, skip loci where the fraction of samples covered by reads is below FLOAT. [0]
+.TP
+.B -e
+Perform max-likelihood inference only, including estimating the site allele frequency,
+testing Hardy-Weinberg equlibrium and testing associations with LRT.
+.TP
+.B -g
+Call per-sample genotypes at variant sites (force -c)
+.TP
+.BI -i \ FLOAT
+Ratio of INDEL-to-SNP mutation rate [0.15]
+.TP
+.BI -p \ FLOAT
+A site is considered to be a variant if P(ref|D)<FLOAT [0.5]
+.TP
+.BI -P \ STR
+Prior or initial allele frequency spectrum. If STR can be
+.IR full ,
+.IR cond2 ,
+.I flat
+or the file consisting of error output from a previous variant calling
+run.
+.TP
+.BI -t \ FLOAT
+Scaled muttion rate for variant calling [0.001]
+.TP
+.BI -T \ STR
+Enable pair/trio calling. For trio calling, option
+.B -s
+is usually needed to be applied to configure the trio members and their ordering.
+In the file supplied to the option
+.BR -s ,
+the first sample must be the child, the second the father and the third the mother.
+The valid values of
+.I STR
+are `pair', `trioauto', `trioxd' and `trioxs', where `pair' calls differences between two input samples, and `trioxd' (`trioxs') specifies that the input
+is from the X chromosome non-PAR regions and the child is a female (male). [null]
+.TP
+.B -v
+Output variant sites only (force -c)
+.TP
+.B Contrast Calling and Association Test Options:
+.TP
+.BI -1 \ INT
+Number of group-1 samples. This option is used for dividing the samples into
+two groups for contrast SNP calling or association test.
+When this option is in use, the following VCF INFO will be outputted:
+PC2, PCHI2 and QCHI2. [0]
+.TP
+.BI -U \ INT
+Number of permutations for association test (effective only with
+.BR -1 )
+[0]
+.TP
+.BI -X \ FLOAT
+Only perform permutations for P(chi^2)<FLOAT (effective only with
+.BR -U )
+[0.01]
+.RE
+
+.TP
+.B index
+.B bcftools index
+.I in.bcf
+
+Index sorted BCF for random access.
+.RE
+
+.TP
+.B cat
+.B bcftools cat
+.I in1.bcf
+.RI [ "in2.bcf " [ ... "]]]"
+
+Concatenate BCF files. The input files are required to be sorted and
+have identical samples appearing in the same order.
+.RE
.SH SAM FORMAT
-SAM is TAB-delimited. Apart from the header lines, which are started
+Sequence Alignment/Map (SAM) format is TAB-delimited. Apart from the header lines, which are started
with the `@' symbol, each alignment line consists of:
.TS
n | l | l .
Col Field Description
_
-1 QNAME Query (pair) NAME
+1 QNAME Query template/pair NAME
2 FLAG bitwise FLAG
3 RNAME Reference sequence NAME
4 POS 1-based leftmost POSition/coordinate of clipped sequence
6 CIAGR extended CIGAR string
7 MRNM Mate Reference sequence NaMe (`=' if same as RNAME)
8 MPOS 1-based Mate POSistion
-9 ISIZE Inferred insert SIZE
+9 TLEN inferred Template LENgth (insert size)
10 SEQ query SEQuence on the same strand as the reference
11 QUAL query QUALity (ASCII-33 gives the Phred base quality)
-12 OPT variable OPTional fields in the format TAG:VTYPE:VALUE
+12+ OPT variable OPTional fields in the format TAG:VTYPE:VALUE
.TE
.PP
0x0400 d the read is either a PCR or an optical duplicate
.TE
+where the second column gives the string representation of the FLAG field.
+
+.SH VCF FORMAT
+
+The Variant Call Format (VCF) is a TAB-delimited format with each data line consists of the following fields:
+.TS
+center box;
+cb | cb | cb
+n | l | l .
+Col Field Description
+_
+1 CHROM CHROMosome name
+2 POS the left-most POSition of the variant
+3 ID unique variant IDentifier
+4 REF the REFerence allele
+5 ALT the ALTernate allele(s), separated by comma
+6 QUAL variant/reference QUALity
+7 FILTER FILTers applied
+8 INFO INFOrmation related to the variant, separated by semi-colon
+9 FORMAT FORMAT of the genotype fields, separated by colon (optional)
+10+ SAMPLE SAMPLE genotypes and per-sample information (optional)
+.TE
+
+.PP
+The following table gives the
+.B INFO
+tags used by samtools and bcftools.
+
+.TS
+center box;
+cb | cb | cb
+l | l | l .
+Tag Format Description
+_
+AF1 double Max-likelihood estimate of the site allele frequency (AF) of the first ALT allele
+DP int Raw read depth (without quality filtering)
+DP4 int[4] # high-quality reference forward bases, ref reverse, alternate for and alt rev bases
+FQ int Consensus quality. Positive: sample genotypes different; negative: otherwise
+MQ int Root-Mean-Square mapping quality of covering reads
+PC2 int[2] Phred probability of AF in group1 samples being larger (,smaller) than in group2
+PCHI2 double Posterior weighted chi^2 P-value between group1 and group2 samples
+PV4 double[4] P-value for strand bias, baseQ bias, mapQ bias and tail distance bias
+QCHI2 int Phred-scaled PCHI2
+RP int # permutations yielding a smaller PCHI2
+CLR int Phred log ratio of genotype likelihoods with and without the trio/pair constraint
+UGT string Most probable genotype configuration without the trio constraint
+CGT string Most probable configuration with the trio constraint
+.TE
+
.SH EXAMPLES
.IP o 2
Import SAM to BAM when
.IR RG:Z:454 .
.IP o 2
-Call SNPs and short indels for one diploid individual:
+Call SNPs and short INDELs for one diploid individual:
- samtools pileup -vcf ref.fa aln.bam > var.raw.plp
- samtools.pl varFilter -D 100 var.raw.plp > var.flt.plp
- awk '($3=="*"&&$6>=50)||($3!="*"&&$6>=20)' var.flt.plp > var.final.plp
+ samtools mpileup -ugf ref.fa aln.bam | bcftools view -bvcg - > var.raw.bcf
+ bcftools view var.raw.bcf | vcfutils.pl varFilter -D 100 > var.flt.vcf
The
.B -D
adjusted to about twice the average read depth. One may consider to add
.B -C50
to
-.B pileup
+.B mpileup
if mapping quality is overestimated for reads containing excessive
mismatches. Applying this option usually helps
.B BWA-short
-but may not other mappers. It also potentially increases reference
-biases.
+but may not other mappers.
+
+.IP o 2
+Generate the consensus sequence for one diploid individual:
+
+ samtools mpileup -uf ref.fa aln.bam | bcftools view -cg - | vcfutils.pl vcf2fq > cns.fq
+
+.IP o 2
+Call somatic mutations from a pair of samples:
+
+ samtools mpileup -DSuf ref.fa aln.bam | bcftools view -bvcgT pair - > var.bcf
+
+In the output INFO field,
+.I CLR
+gives the Phred-log ratio between the likelihood by treating the
+two samples independently, and the likelihood by requiring the genotype to be identical.
+This
+.I CLR
+is effectively a score measuring the confidence of somatic calls. The higher the better.
+
+.IP o 2
+Call de novo and somatic mutations from a family trio:
+
+ samtools mpileup -DSuf ref.fa aln.bam | bcftools view -bvcgT pair -s samples.txt - > var.bcf
+
+File
+.I samples.txt
+should consist of three lines specifying the member and order of samples (in the order of child-father-mother).
+Similarly,
+.I CLR
+gives the Phred-log likelihood ratio with and without the trio constraint.
+.I UGT
+shows the most likely genotype configuration without the trio constraint, and
+.I CGT
+gives the most likely genotype configuration satisfying the trio constraint.
.IP o 2
-Call SNPs (not short indels) for multiple diploid individuals:
+Phase one individual:
- samtools mpileup -augf ref.fa *.bam | bcftools view -bcv - > snp.raw.bcf
- bcftools view snp.raw.bcf | vcfutils.pl filter4vcf -D 2000 | bgzip > snp.flt.vcf.gz
+ samtools calmd -AEur aln.bam ref.fa | samtools phase -b prefix - > phase.out
+
+The
+.B calmd
+command is used to reduce false heterozygotes around INDELs.
+
+.IP o 2
+Call SNPs and short indels for multiple diploid individuals:
+
+ samtools mpileup -P ILLUMINA -ugf ref.fa *.bam | bcftools view -bcvg - > var.raw.bcf
+ bcftools view var.raw.bcf | vcfutils.pl varFilter -D 2000 > var.flt.vcf
Individuals are identified from the
.B SM
tags in the
.B @RG
header lines. Individuals can be pooled in one alignment file; one
-individual can also be separated into multiple files. Similarly, one may
-consider to apply
-.B -C50
-to
-.BR mpileup .
-SNP calling in this way also works for single sample and has the
-advantage of enabling more powerful filtering. The drawback is the lack
-of short indel calling, which may be implemented in future.
+individual can also be separated into multiple files. The
+.B -P
+option specifies that indel candidates should be collected only from
+read groups with the
+.B @RG-PL
+tag set to
+.IR ILLUMINA .
+Collecting indel candidates from reads sequenced by an indel-prone
+technology may affect the performance of indel calling.
.IP o 2
Derive the allele frequency spectrum (AFS) on a list of sites from multiple individuals:
- samtools mpileup -gf ref.fa *.bam > all.bcf
+ samtools mpileup -Igf ref.fa *.bam > all.bcf
bcftools view -bl sites.list all.bcf > sites.bcf
bcftools view -cGP cond2 sites.bcf > /dev/null 2> sites.1.afs
bcftools view -cGP sites.1.afs sites.bcf > /dev/null 2> sites.2.afs
.IP o 2
Dump BAQ applied alignment for other SNP callers:
- samtools calmd -br aln.bam > aln.baq.bam
+ samtools calmd -bAr aln.bam > aln.baq.bam
It adds and corrects the
.B NM
.B calmd
command also comes with the
.B -C
-option, the same as the on in
+option, the same as the one in
.B pileup
and
.BR mpileup .
.IP o 2
Unaligned words used in bam_import.c, bam_endian.h, bam.c and bam_aux.c.
.IP o 2
-In merging, the input files are required to have the same number of
-reference sequences. The requirement can be relaxed. In addition,
-merging does not reconstruct the header dictionaries
-automatically. Endusers have to provide the correct header. Picard is
-better at merging.
-.IP o 2
Samtools paired-end rmdup does not work for unpaired reads (e.g. orphan
reads or ends mapped to different chromosomes). If this is a concern,
please use Picard's MarkDuplicate which correctly handles these cases,
.PP
Heng Li from the Sanger Institute wrote the C version of samtools. Bob
Handsaker from the Broad Institute implemented the BGZF library and Jue
-Ruan from Beijing Genomics Institute wrote the RAZF library. Various
-people in the 1000 Genomes Project contributed to the SAM format
+Ruan from Beijing Genomics Institute wrote the RAZF library. John
+Marshall and Petr Danecek contribute to the source code and various
+people from the 1000 Genomes Project have contributed to the SAM format
specification.
.SH SEE ALSO
projects / samtools.git / blobdiff