Merge commit 'upstream/0.1.10'

[samtools.git] / samtools.txt

diff --git a/samtools.txt b/samtools.txt
index 63e6a25ec72823931cda29bd2ffd9ddad745b2db..41cabe5053986a44f415ae35981c0f61738a7996 100644 (file)
--- a/samtools.txt
+++ b/samtools.txt
@@ -12,81 +12,222 @@ SYNOPSIS
 
        samtools index aln.sorted.bam
 
 
        samtools index aln.sorted.bam
 

+       samtools idxstats aln.sorted.bam
+

        samtools view aln.sorted.bam chr2:20,100,000-20,200,000
 
        samtools merge out.bam in1.bam in2.bam in3.bam
 
        samtools faidx ref.fasta
 
        samtools view aln.sorted.bam chr2:20,100,000-20,200,000
 
        samtools merge out.bam in1.bam in2.bam in3.bam
 
        samtools faidx ref.fasta
 

-       samtools pileup -f ref.fasta aln.sorted.bam
+       samtools pileup -vcf ref.fasta aln.sorted.bam
+
+       samtools mpileup -C50 -gf ref.fasta -r chr3:1,000-2,000 in1.bam in2.bam

 
        samtools tview aln.sorted.bam ref.fasta
 
 
 DESCRIPTION
 
        samtools tview aln.sorted.bam ref.fasta
 
 
 DESCRIPTION

-       Samtools  is  a  set of utilities that manipulate alignments in the BAM
+       Samtools is a set of utilities that manipulate alignments  in  the  BAM

        format. It imports from and exports to the SAM (Sequence Alignment/Map)
        format. It imports from and exports to the SAM (Sequence Alignment/Map)

-       format,  does  sorting,  merging  and  indexing, and allows to retrieve
+       format, does sorting, merging and  indexing,  and  allows  to  retrieve

        reads in any regions swiftly.
 
        reads in any regions swiftly.
 

-       Samtools is designed to work on a stream. It regards an input file  `-'
-       as  the  standard  input (stdin) and an output file `-' as the standard
+       Samtools  is designed to work on a stream. It regards an input file `-'
+       as the standard input (stdin) and an output file `-'  as  the  standard

        output (stdout). Several commands can thus be combined with Unix pipes.
        Samtools always output warning and error messages to the standard error
        output (stderr).
 
        output (stdout). Several commands can thus be combined with Unix pipes.
        Samtools always output warning and error messages to the standard error
        output (stderr).
 

-       Samtools is also able to open a BAM (not SAM) file on a remote  FTP  or
-       HTTP  server  if  the  BAM file name starts with `ftp://' or `http://'.
-       Samtools checks the current working directory for the  index  file  and
-       will  download  the  index upon absence. Samtools does not retrieve the
+       Samtools  is  also able to open a BAM (not SAM) file on a remote FTP or
+       HTTP server if the BAM file name starts  with  `ftp://'  or  `http://'.
+       Samtools  checks  the  current working directory for the index file and
+       will download the index upon absence. Samtools does  not  retrieve  the

        entire alignment file unless it is asked to do so.
 
 
 COMMANDS AND OPTIONS
        entire alignment file unless it is asked to do so.
 
 
 COMMANDS AND OPTIONS

-       import    samtools import <in.ref_list> <in.sam> <out.bam>
+       view      samtools  view  [-bchuHS]  [-t  in.refList]  [-o  output] [-f
+                 reqFlag] [-F skipFlag] [-q minMapQ] [-l  library]  [-r  read-
+                 Group] [-R rgFile] <in.bam>|<in.sam> [region1 [...]]
+
+                 Extract/print  all or sub alignments in SAM or BAM format. If
+                 no region is specified, all the alignments will  be  printed;
+                 otherwise  only  alignments overlapping the specified regions
+                 will be output. An alignment may be given multiple  times  if
+                 it is overlapping several regions. A region can be presented,
+                 for example, in  the  following  format:  `chr2'  (the  whole
+                 chr2),  `chr2:1000000'  (region starting from 1,000,000bp) or
+                 `chr2:1,000,000-2,000,000'  (region  between  1,000,000   and
+                 2,000,000bp  including  the  end  points).  The coordinate is
+                 1-based.
+
+                 OPTIONS:
+
+                 -b      Output in the BAM format.
+
+                 -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]
+
+                 -F INT  Skip alignments with bits present in INT [0]
+
+                 -h      Include the header in the output.
+
+                 -H      Output the header only.
+
+                 -l STR  Only output reads in library STR [null]
+
+                 -o FILE Output file [stdout]
+
+                 -q INT  Skip alignments with MAPQ smaller than INT [0]
+
+                 -r STR  Only output reads in read group STR [null]
+
+                 -R FILE Output reads in read groups listed in FILE [null]
+
+                 -S      Input is in SAM. If @SQ header lines are absent,  the
+                         `-t' option is required.
+
+                 -c      Instead  of  printing the alignments, only count them
+                         and print the total number. All filter options,  such
+                         as `-f', `-F' and `-q' , are taken into account.
+
+                 -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  reference  sequences  in sorting. If you run
+                         `samtools faidx <ref.fa>', the resultant  index  file
+                         <ref.fa>.fai  can be used as this <in.ref_list> file.

 
 

-                 Since 0.1.4, this command is an alias of:
+                 -u      Output uncompressed BAM. This option saves time spent
+                         on  compression/decomprssion  and  is  thus preferred
+                         when the output is piped to another samtools command.

 
 

-                 samtools view -bt <in.ref_list> -o <out.bam> <in.sam>
+
+       tview     samtools tview <in.sorted.bam> [ref.fasta]
+
+                 Text  alignment viewer (based on the ncurses library). In the
+                 viewer, press `?' for help and press `g' to check the  align-
+                 ment    start    from   a   region   in   the   format   like
+                 `chr10:10,000,000' or `=10,000,000'  when  viewing  the  same
+                 reference sequence.

 
 
 
 

-       sort      samtools sort [-n] [-m maxMem] <in.bam> <out.prefix>
+       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. Align-
+                 ment records are grouped by sample identifiers in @RG  header
+                 lines.  If  sample identifiers are absent, each input file is
+                 regarded as one sample.
+
+                 OPTIONS:
+
+                 -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.
+
+                 -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 for
+                         BWA is 50. [0]
+
+                 -e INT  Phred-scaled gap extension sequencing error probabil-
+                         ity. Reducing INT leads to longer indels. [20]
+
+                 -f FILE The reference file [null]
+
+                 -g      Compute  genotype  likelihoods and output them in the
+                         binary call format (BCF).
+
+                 -h INT  Coefficient for modeling homopolymer errors. Given an
+                         l-long  homopolymer  run,  the sequencing error of an
+                         indel of size s is modeled as INT*s/l.  [100]
+
+                 -l FILE File containing a list of sites where pileup  or  BCF
+                         is outputted [null]
+
+                 -o INT  Phred-scaled  gap  open sequencing error probability.
+                         Reducing INT leads to more indel calls. [40]
+
+                 -P STR  Comma dilimited list of platforms (determined by @RG-
+                         PL)  from  which indel candidates are obtained. It is
+                         recommended to collect indel candidates from sequenc-
+                         ing  technologies that have low indel error rate such
+                         as ILLUMINA. [all]
+
+                 -q INT  Minimum mapping quality for an alignment to  be  used
+                         [0]
+
+                 -Q INT  Minimum base quality for a base to be considered [13]
+
+                 -r STR  Only generate pileup in region STR [all sites]
+
+                 -u      Similar to -g except that the output is  uncompressed
+                         BCF, which is preferred for piping.
+
+
+       reheader  samtools reheader <in.header.sam> <in.bam>
+
+                 Replace   the   header   in   in.bam   with   the  header  in
+                 in.header.sam.  This command is much  faster  than  replacing
+                 the header with a BAM->SAM->BAM conversion.
+
+
+       sort      samtools sort [-no] [-m maxMem] <in.bam> <out.prefix>

 
                  Sort  alignments  by  leftmost  coordinates.  File  <out.pre-
                  fix>.bam will be created. This command may also create tempo-
 
                  Sort  alignments  by  leftmost  coordinates.  File  <out.pre-
                  fix>.bam will be created. This command may also create tempo-

-                 rary files <out.prefix>.%d.bam when the whole alignment  can-
+                 rary  files <out.prefix>.%d.bam when the whole alignment can-

                  not be fitted into memory (controlled by option -m).
 
                  OPTIONS:
 
                  not be fitted into memory (controlled by option -m).
 
                  OPTIONS:
 

+                 -o      Output the final alignment to the standard output.
+

                  -n      Sort by read names rather than by chromosomal coordi-
                          nates
 
                  -n      Sort by read names rather than by chromosomal coordi-
                          nates
 

-                 -m INT  Approximately   the    maximum    required    memory.
+                 -m INT  Approximately    the    maximum    required   memory.

                          [500000000]
 
 
                          [500000000]
 
 

-       merge     samtools   merge   [-h   inh.sam]  [-n]  <out.bam>  <in1.bam>
-                 <in2.bam> [...]
+       merge     samtools  merge  [-nur]  [-h  inh.sam]  [-R  reg]   <out.bam>
+                 <in1.bam> <in2.bam> [...]

 
                  Merge multiple sorted alignments.  The header reference lists
 
                  Merge multiple sorted alignments.  The header reference lists

-                 of  all  the input BAM files, and the @SQ headers of inh.sam,
+                 of all the input BAM files, and the @SQ headers  of  inh.sam,

                  if  any,  must  all  refer  to  the  same  set  of  reference
                  if  any,  must  all  refer  to  the  same  set  of  reference

-                 sequences.   The header reference list and (unless overridden
-                 by -h) `@' headers of in1.bam will be copied to out.bam,  and
+                 sequences.  The header reference list and (unless  overridden
+                 by  -h) `@' headers of in1.bam will be copied to out.bam, and

                  the headers of other files will be ignored.
 
                  OPTIONS:
 
                  the headers of other files will be ignored.
 
                  OPTIONS:
 

-                 -h FILE Use  the lines of FILE as `@' headers to be copied to
+                 -h FILE Use the lines of FILE as `@' headers to be copied  to

                          out.bam, replacing any header lines that would other-
                          out.bam, replacing any header lines that would other-

-                         wise  be  copied  from in1.bam.  (FILE is actually in
-                         SAM format, though any alignment records it may  con-
+                         wise be copied from in1.bam.  (FILE  is  actually  in
+                         SAM  format, though any alignment records it may con-

                          tain are ignored.)
 
                          tain are ignored.)
 

+                 -R STR  Merge files in the specified region indicated by STR
+
+                 -r      Attach an RG tag to each alignment. The tag value  is
+                         inferred from file names.
+

                  -n      The  input alignments are sorted by read names rather
                          than by chromosomal coordinates
 
                  -n      The  input alignments are sorted by read names rather
                          than by chromosomal coordinates
 

+                 -u      Uncompressed BAM output
+

 
        index     samtools index <aln.bam>
 
 
        index     samtools index <aln.bam>
 


@@ -94,70 +235,78 @@ COMMANDS AND OPTIONS
                  <aln.bam>.bai will be created.
 
 
                  <aln.bam>.bai will be created.
 
 

-       view      samtools  view  [-bhuHS]  [-t  in.refList]  [-o  output]  [-f
-                 reqFlag] [-F skipFlag] [-q minMapQ] [-l  library]  [-r  read-
-                 Group] <in.bam>|<in.sam> [region1 [...]]
+       idxstats  samtools idxstats <aln.bam>

 
 

-                 Extract/print  all or sub alignments in SAM or BAM format. If
-                 no region is specified, all the alignments will  be  printed;
-                 otherwise  only  alignments overlapping the specified regions
-                 will be output. An alignment may be given multiple  times  if
-                 it is overlapping several regions. A region can be presented,
-                 for example, in the following format: `chr2',  `chr2:1000000'
-                 or `chr2:1,000,000-2,000,000'. The coordinate is 1-based.
+                 Retrieve and print stats in the index file. The output is TAB
+                 delimited with each line  consisting  of  reference  sequence
+                 name, sequence length, # mapped reads and # unmapped reads.

 
 

-                 OPTIONS:

 
 

-                 -b      Output in the BAM format.
+       faidx     samtools faidx <ref.fasta> [region1 [...]]

 
 

-                 -u      Output uncompressed BAM. This option saves time spent
-                         on compression/decomprssion  and  is  thus  preferred
-                         when the output is piped to another samtools command.
+                 Index  reference sequence in the FASTA format or extract sub-
+                 sequence from indexed reference sequence.  If  no  region  is
+                 specified,   faidx   will   index   the   file   and   create
+                 <ref.fasta>.fai on the disk. If regions are speficified,  the
+                 subsequences  will  be retrieved and printed to stdout in the
+                 FASTA format. The input file can be compressed  in  the  RAZF
+                 format.

 
 

-                 -h      Include the header in the output.

 
 

-                 -H      Output the header only.
+       fixmate   samtools fixmate <in.nameSrt.bam> <out.bam>

 
 

-                 -S      Input is in SAM. If @SQ header lines are absent,  the
-                         `-t' option is required.
+                 Fill in mate coordinates, ISIZE and mate related flags from a
+                 name-sorted alignment.

 
 

-                 -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  reference  sequences  in sorting. If you run
-                         `samtools faidx <ref.fa>', the resultant  index  file
-                         <ref.fa>.fai  can be used as this <in.ref_list> file.

 
 

-                 -o FILE Output file [stdout]
+       rmdup     samtools rmdup [-sS] <input.srt.bam> <out.bam>

 
 

-                 -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]
+                 Remove potential PCR duplicates: if multiple read pairs  have
+                 identical  external  coordinates,  only  retain the pair with
+                 highest mapping quality.  In the paired-end mode,  this  com-
+                 mand  ONLY  works  with  FR orientation and requires ISIZE is
+                 correctly set. It does not work for unpaired reads (e.g.  two
+                 ends mapped to different chromosomes or orphan reads).

 
 

-                 -F INT  Skip alignments with bits present in INT [0]
+                 OPTIONS:

 
 

-                 -q INT  Skip alignments with MAPQ smaller than INT [0]
+                 -s      Remove  duplicate  for  single-end reads. By default,
+                         the command works for paired-end reads only.

 
 

-                 -l STR  Only output reads in library STR [null]
+                 -S      Treat paired-end reads and single-end reads.

 
 

-                 -r STR  Only output reads in read group STR [null]

 
 

+       calmd     samtools calmd [-eubSr] [-C capQcoef] <aln.bam> <ref.fasta>

 
 

-       faidx     samtools faidx <ref.fasta> [region1 [...]]
+                 Generate the MD tag. If the MD tag is already  present,  this
+                 command  will  give a warning if the MD tag generated is dif-
+                 ferent from the existing tag. Output SAM by default.

 
 

-                 Index reference sequence in the FASTA format or extract  sub-
-                 sequence  from  indexed  reference  sequence. If no region is
-                 specified,   faidx   will   index   the   file   and   create
-                 <ref.fasta>.fai  on the disk. If regions are speficified, the
-                 subsequences will be retrieved and printed to stdout  in  the
-                 FASTA  format.  The  input file can be compressed in the RAZF
-                 format.
+                 OPTIONS:

 
 

+                 -A      When used jointly with -r this option overwrites  the
+                         original base quality.

 
 

-       pileup    samtools  pileup  [-f  in.ref.fasta]  [-t  in.ref_list]   [-l
-                 in.site_list]    [-iscgS2]   [-T   theta]   [-N   nHap]   [-r
-                 pairDiffRate] <in.bam>|<in.sam>
+                 -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.
+
+                 -u      Output uncompressed BAM
+
+                 -b      Output compressed BAM
+
+                 -S      The input is SAM with header lines
+
+                 -C INT  Coefficient  to  cap mapping quality of poorly mapped
+                         reads. See the pileup command for details. [0]
+
+                 -r      Compute the BQ tag without changing the base quality.
+
+
+       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>

 
                  Print the alignment in the pileup format. In the pileup  for-
                  mat,  each  line represents a genomic position, consisting of
 
                  Print the alignment in the pileup format. In the pileup  for-
                  mat,  each  line represents a genomic position, consisting of


@@ -167,25 +316,25 @@ COMMANDS AND OPTIONS
                  end  of  a  read  are all encoded at the read base column. At
                  this column, a dot stands for a match to the  reference  base
                  on  the  forward  strand,  a comma for a match on the reverse
                  end  of  a  read  are all encoded at the read base column. At
                  this column, a dot stands for a match to the  reference  base
                  on  the  forward  strand,  a comma for a match on the reverse

-                 strand, `ACGTN' for a mismatch  on  the  forward  strand  and
-                 `acgtn'  for  a  mismatch  on  the  reverse strand. A pattern
-                 `\+[0-9]+[ACGTNacgtn]+'  indicates  there  is  an   insertion
-                 between  this reference position and the next reference posi-
-                 tion. The length of the insertion is given by the integer  in
-                 the  pattern, followed by the inserted sequence. Similarly, a
-                 pattern `-[0-9]+[ACGTNacgtn]+' represents a deletion from the
-                 reference.  The deleted bases will be presented as `*' in the
-                 following lines. Also at the read base column, a  symbol  `^'
-                 marks  the start of a read segment which is a contiguous sub-
-                 sequence on the read separated by `N/S/H'  CIGAR  operations.
-                 The  ASCII  of the character following `^' minus 33 gives the
-                 mapping quality. A symbol `$' marks the end of  a  read  seg-
-                 ment.
-
-                 If  option -c is applied, the consensus base, consensus qual-
-                 ity, SNP quality and RMS mapping quality of the reads  cover-
-                 ing  the  site  will be inserted between the `reference base'
-                 and the `read bases' columns. An indel occupies an additional
+                 strand, a '>' or '<' for a reference skip, `ACGTN' for a mis-
+                 match on the forward strand and `acgtn' for a mismatch on the
+                 reverse strand. A pattern  `\+[0-9]+[ACGTNacgtn]+'  indicates
+                 there is an insertion between this reference position and the
+                 next reference position. The length of the insertion is given
+                 by  the  integer  in  the  pattern,  followed by the inserted
+                 sequence. Similarly, a pattern `-[0-9]+[ACGTNacgtn]+'  repre-
+                 sents  a  deletion from the reference. The deleted bases will
+                 be presented as `*' in the following lines. Also at the  read
+                 base  column,  a  symbol  `^'  marks the start of a read. The
+                 ASCII of the character following `^' minus 33 gives the  map-
+                 ping quality. A symbol `$' marks the end of a read segment.
+
+                 If  option  -c  is  applied, the consensus base, Phred-scaled
+                 consensus quality, SNP quality (i.e. the Phred-scaled  proba-
+                 bility 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, coordi-
                  nate, a star, the genotype, consensus quality,  SNP  quality,
                  RMS mapping quality, # covering reads, the first alllele, the
                  line.  Each  indel  line consists of chromosome name, coordi-
                  nate, a star, the genotype, consensus quality,  SNP  quality,
                  RMS mapping quality, # covering reads, the first alllele, the


@@ -193,175 +342,212 @@ COMMANDS AND OPTIONS
                  supporting  the  second  allele and # reads containing indels
                  different from the top two alleles.
 
                  supporting  the  second  allele and # reads containing indels
                  different from the top two alleles.
 

-                 OPTIONS:
-
+                 NOTE: Since 0.1.10, the `pileup'  command  is  deprecated  by
+                 `mpileup'.

 
 

-                 -s        Print the mapping quality as the last column.  This
-                           option  makes  the output easier to parse, although
-                           this format is not space efficient.
+                 OPTIONS:

 
 

+                 -B        Disable  the  BAQ computation. See the mpileup com-
+                           mand for details.

 
 

-                 -S        The input file is in SAM.
+                 -c        Call the consensus sequence. Options -T, -N, -I and
+                           -r are only effective when -c or -g is in use.

 
 

+                 -C INT    Coefficient  for downgrading the mapping quality of
+                           poorly mapped reads. See the  mpileup  command  for
+                           details. [0]

 
 

-                 -i        Only output pileup lines containing indels.
-
+                 -d INT    Use  the  first  NUM  reads in the pileup for indel
+                           calling for speed up. Zero for unlimited. [1024]

 
                  -f FILE   The reference sequence in the FASTA  format.  Index
                            file FILE.fai will be created if absent.
 
 
                  -f FILE   The reference sequence in the FASTA  format.  Index
                            file FILE.fai will be created if absent.
 

+                 -g        Generate  genotype  likelihood  in the binary GLFv3
+                           format. This option suppresses -c, -i and -s.  This
+                           option is deprecated by the mpileup command.

 
 

-                 -M INT    Cap mapping quality at INT [60]
+                 -i        Only output pileup lines containing indels.

 
 

+                 -I INT    Phred  probability  of an indel in sequencing/prep.
+                           [40]

 
 

-                 -t FILE   List  of  reference  names ane sequence lengths, in
-                           the format described for  the  import  command.  If
-                           this  option is present, samtools assumes the input
-                           <in.alignment>  is  in  SAM  format;  otherwise  it
-                           assumes in BAM format.
+                 -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

 
 

+                 -m INT    Filter reads  with  flag  containing  bits  in  INT
+                           [1796]

 
 

-                 -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 -s together with -l as in the default
-                           format we may not know the mapping quality.
+                 -M INT    Cap mapping quality at INT [60]

 
 

+                 -N INT    Number of haplotypes in the sample (>=2) [2]

 
 

-                 -c        Call the consensus  sequence  using  MAQ  consensus
-                           model. Options -T, -N, -I and -r are only effective
-                           when -c or -g is in use.
+                 -r FLOAT  Expected  fraction of differences between a pair of
+                           haplotypes [0.001]

 
 

+                 -s        Print the mapping quality as the last column.  This
+                           option  makes  the output easier to parse, although
+                           this format is not space efficient.

 
 

-                 -g        Generate genotype likelihood in  the  binary  GLFv3
-                           format. This option suppresses -c, -i and -s.
+                 -S        The input file is in SAM.

 
 

+                 -t FILE   List of reference names ane  sequence  lengths,  in
+                           the  format  described  for  the import command. If
+                           this option is present, samtools assumes the  input
+                           <in.alignment>  is  in  SAM  format;  otherwise  it
+                           assumes in BAM format.  -s together with -l  as  in
+                           the  default  format  we  may  not know the mapping
+                           quality.

 
 

-                 -T FLOAT  The  theta parameter (error dependency coefficient)
+                 -T FLOAT  The theta parameter (error dependency  coefficient)

                            in the maq consensus calling model [0.85]
 
 
                            in the maq consensus calling model [0.85]
 
 

-                 -N INT    Number of haplotypes in the sample (>=2) [2]
-
-
-                 -r FLOAT  Expected fraction of differences between a pair  of
-                           haplotypes [0.001]
+SAM FORMAT
+       SAM  is  TAB-delimited.  Apart from the header lines, which are started
+       with the `@' symbol, each alignment line consists of:

 
 
 
 

-                 -I INT    Phred  probability  of an indel in sequencing/prep.
-                           [40]
+       +----+-------+----------------------------------------------------------+
+       |Col | Field |                       Description                        |
+       +----+-------+----------------------------------------------------------+
+       | 1  | QNAME | Query (pair) NAME                                        |
+       | 2  | FLAG  | bitwise FLAG                                             |
+       | 3  | RNAME | Reference sequence NAME                                  |
+       | 4  | POS   | 1-based leftmost POSition/coordinate of clipped sequence |
+       | 5  | MAPQ  | MAPping Quality (Phred-scaled)                           |
+       | 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                                     |
+       |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   |
+       +----+-------+----------------------------------------------------------+

 
 

+       Each bit in the FLAG field is defined as:

 
 
 
 

-       tview     samtools tview <in.sorted.bam> [ref.fasta]
+          +-------+-----+--------------------------------------------------+
+          | Flag  | Chr |                   Description                    |
+          +-------+-----+--------------------------------------------------+
+          |0x0001 |  p  | the read is paired in sequencing                 |
+          |0x0002 |  P  | the read is mapped in a proper pair              |
+          |0x0004 |  u  | the query sequence itself is unmapped            |
+          |0x0008 |  U  | the mate is unmapped                             |
+          |0x0010 |  r  | strand of the query (1 for reverse)              |
+          |0x0020 |  R  | strand of the mate                               |
+          |0x0040 |  1  | the read is the first read in a pair             |
+          |0x0080 |  2  | the read is the second read in a pair            |
+          |0x0100 |  s  | the alignment is not primary                     |
+          |0x0200 |  f  | the read fails platform/vendor quality checks    |
+          |0x0400 |  d  | the read is either a PCR or an optical duplicate |
+          +-------+-----+--------------------------------------------------+

 
 

-                 Text alignment viewer (based on the ncurses library). In  the
-                 viewer,  press `?' for help and press `g' to check the align-
-                 ment   start   from   a   region   in   the    format    like
-                 `chr10:10,000,000'.
+EXAMPLES
+       o Import SAM to BAM when @SQ lines are present in the header:

 
 

+           samtools view -bS aln.sam > aln.bam

 
 

+         If @SQ lines are absent:

 
 

-       fixmate   samtools fixmate <in.nameSrt.bam> <out.bam>
+           samtools faidx ref.fa
+           samtools view -bt ref.fa.fai aln.sam > aln.bam

 
 

-                 Fill in mate coordinates, ISIZE and mate related flags from a
-                 name-sorted alignment.
+         where ref.fa.fai is generated automatically by the faidx command.

 
 
 
 

-       rmdup     samtools rmdup <input.srt.bam> <out.bam>
+       o Attach the RG tag while merging sorted alignments:

 
 

-                 Remove potential PCR duplicates: if multiple read pairs  have
-                 identical  external  coordinates,  only  retain the pair with
-                 highest mapping quality.  This command  ONLY  works  with  FR
-                 orientation and requires ISIZE is correctly set.
+           perl      -e       'print       "@RG\tID:ga\tSM:hs\tLB:ga\tPL:Illu-
+         mina\n@RG\tID:454\tSM:hs\tLB:454\tPL:454\n"' > rg.txt
+           samtools merge -rh rg.txt merged.bam ga.bam 454.bam

 
 

+         The value in a RG tag is determined by the file name the read is com-
+         ing from. In this example, in the merged.bam, reads from ga.bam  will
+         be  attached  RG:Z:ga,  while  reads  from  454.bam  will be attached
+         RG:Z:454.

 
 
 
 

-       rmdupse   samtools rmdupse <input.srt.bam> <out.bam>
+       o Call SNPs and short indels for one diploid individual:

 
 

-                 Remove potential duplicates for single-ended reads. This com-
-                 mand will treat all reads as single-ended even  if  they  are
-                 paired in fact.
+           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 -D option of varFilter controls the  maximum  read  depth,  which
+         should  be  adjusted  to about twice the average read depth.  One may
+         consider to add -C50 to mpileup if mapping quality  is  overestimated
+         for  reads containing excessive mismatches. Applying this option usu-
+         ally helps BWA-short but may not other mappers.

 
 
 
 

-       fillmd    samtools fillmd [-e] <aln.bam> <ref.fasta>
+       o Call SNPs and short indels for multiple diploid individuals:

 
 

-                 Generate  the  MD tag. If the MD tag is already present, this
-                 command will give a warning if the MD tag generated  is  dif-
-                 ferent from the existing tag.
+           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

 
 

-                 OPTIONS:
+         Individuals are identified from the SM tags in the @RG header  lines.
+         Individuals  can  be pooled in one alignment file; one individual can
+         also be separated into multiple files. The -P option  specifies  that
+         indel  candidates  should be collected only from read groups with the
+         @RG-PL tag set to ILLUMINA.  Collecting indel candidates  from  reads
+         sequenced  by an indel-prone technology may affect the performance of
+         indel calling.

 
 

-                 -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.

 
 

+       o Derive the allele frequency spectrum (AFS) on a list  of  sites  from
+         multiple individuals:

 
 

+           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
+           bcftools view -cGP sites.2.afs sites.bcf > /dev/null 2> sites.3.afs
+           ......

 
 

-SAM FORMAT
-       SAM  is  TAB-delimited.  Apart from the header lines, which are started
-       with the `@' symbol, each alignment line consists of:
+         where sites.list contains the list of sites with each line consisting
+         of the reference sequence name and position. The  following  bcftools
+         commands estimate AFS by EM.

 
 
 
 

-       +----+-------+----------------------------------------------------------+
-       |Col | Field |                       Description                        |
-       +----+-------+----------------------------------------------------------+
-       | 1  | QNAME | Query (pair) NAME                                        |
-       | 2  | FLAG  | bitwise FLAG                                             |
-       | 3  | RNAME | Reference sequence NAME                                  |
-       | 4  | POS   | 1-based leftmost POSition/coordinate of clipped sequence |
-       | 5  | MAPQ  | MAPping Quality (Phred-scaled)                           |
-       | 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                                     |
-       |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   |
-       +----+-------+----------------------------------------------------------+
+       o Dump BAQ applied alignment for other SNP callers:

 
 

-       Each bit in the FLAG field is defined as:
+           samtools calmd -bAr aln.bam > aln.baq.bam

 
 

+         It  adds  and corrects the NM and MD tags at the same time. The calmd
+         command also comes with the -C option, the same as the one in  pileup
+         and mpileup.  Apply if it helps.

 
 

-             +-------+--------------------------------------------------+
-             | Flag  |                   Description                    |
-             +-------+--------------------------------------------------+
-             |0x0001 | the read is paired in sequencing                 |
-             |0x0002 | the read is mapped in a proper pair              |
-             |0x0004 | the query sequence itself is unmapped            |
-             |0x0008 | the mate is unmapped                             |
-             |0x0010 | strand of the query (1 for reverse)              |
-             |0x0020 | strand of the mate                               |
-             |0x0040 | the read is the first read in a pair             |
-             |0x0080 | the read is the second read in a pair            |
-             |0x0100 | the alignment is not primary                     |
-             |0x0200 | the read fails platform/vendor quality checks    |
-             |0x0400 | the read is either a PCR or an optical duplicate |
-             +-------+--------------------------------------------------+

 
 LIMITATIONS
 
 LIMITATIONS

-       o Unaligned  words  used  in  bam_import.c,  bam_endian.h,  bam.c   and
+       o Unaligned   words  used  in  bam_import.c,  bam_endian.h,  bam.c  and

          bam_aux.c.
 
          bam_aux.c.
 

-       o CIGAR operation P is not properly handled at the moment.
-
-       o 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
+       o 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.
 
          merging.
 

-       o Samtools' rmdup does not work for single-end data and does not remove
-         duplicates across chromosomes. Picard is better.
+       o 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, although a little slower.

 
 
 AUTHOR
 
 
 AUTHOR

-       Heng Li from the Sanger Institute wrote the C version of samtools.  Bob
+       Heng  Li from the Sanger Institute wrote the C version of samtools. Bob

        Handsaker from the Broad Institute implemented the BGZF library and Jue
        Handsaker from the Broad Institute implemented the BGZF library and Jue

-       Ruan from Beijing Genomics Institute wrote the  RAZF  library.  Various
-       people  in the 1000Genomes Project contributed to the SAM format speci-
+       Ruan  from Beijing Genomics Institute wrote the RAZF library. John Mar-
+       shall and Petr Danecek contribute to the source code and various people
+       from the 1000 Genomes Project have contributed to the SAM format speci-

        fication.
 
 
        fication.
 
 


@@ -370,4 +556,4 @@ SEE ALSO
 
 
 
 
 
 

-samtools-0.1.6                 2 September 2009                    samtools(1)
+samtools-0.1.10                15 November 2010                    samtools(1)