sam文件转换为bam文件——SAMtools

在转录组数据与参考基因组进行比对后，得到sam文件，后续分析需要将sam转换为bam，这里用到的工具是SAMtools。

序列比对 —— Hisat2 - (jianshu.com)

SAMtools的主要功能是读取、写出、编辑、查看SAM/BAM/CRAM格式的数据文件。

SAMtools官方网站：
http://www.htslib.org/

1.下载安装

链接：
http://www.htslib.org/download/

$ tar -jxvf samtools-1.14.tar.bz2

安装：

$ cd ~/samtools-1.14
$ ./configure --prefix=/your/path
$ make
$ make install

2. 基础命令

$ samtools

Program: samtools (Tools for alignments in the SAM format)
Version: 1.14 (using htslib 1.14)

Usage:   samtools  [options]

Commands:
  -- Indexing
     dict           create a sequence dictionary file
     faidx          index/extract FASTA
     fqidx          index/extract FASTQ
     index          index alignment

  -- Editing
     calmd          recalculate MD/NM tags and '=' bases
     fixmate        fix mate information
     reheader       replace BAM header
     targetcut      cut fosmid regions (for fosmid pool only)
     addreplacerg   adds or replaces RG tags
     markdup        mark duplicates
     ampliconclip   clip oligos from the end of reads

  -- File operations
     collate        shuffle and group alignments by name
     cat            concatenate BAMs
     merge          merge sorted alignments
     mpileup        multi-way pileup
     sort           sort alignment file
     split          splits a file by read group
     quickcheck     quickly check if SAM/BAM/CRAM file appears intact
     fastq          converts a BAM to a FASTQ
     fasta          converts a BAM to a FASTA
     import         Converts FASTA or FASTQ files to SAM/BAM/CRAM

  -- Statistics
     bedcov         read depth per BED region
     coverage       alignment depth and percent coverage
     depth          compute the depth
     flagstat       simple stats
     idxstats       BAM index stats
     phase          phase heterozygotes
     stats          generate stats (former bamcheck)
     ampliconstats  generate amplicon specific stats

  -- Viewing
     flags          explain BAM flags
     tview          text alignment viewer
     view           SAM<->BAM<->CRAM conversion
     depad          convert padded BAM to unpadded BAM
     samples        list the samples in a set of SAM/BAM/CRAM files

  -- Misc
     help [cmd]     display this help message or help for [cmd]
     version        detailed version information

这里看到基础命令主要分为六个模块，分别为索引、编辑、文件操作、统计、视图以及其他。这里想要SAM格式转为BAM格式，主要用到的是Viewing模块中的view。

3. sam格式转换为bam

说明书：
http://www.htslib.org/workflow/fastq.html

在老版本1.9的samtools中，需要用-s 指定sam文件，1.14中不需要指定。

samtools view命令完成sam转为bam。

3.1 view命令基础功能

$ samtools view

Usage: samtools view [options] || [region ...]

Output options:
  -b, --bam                  Output BAM
  -C, --cram                 Output CRAM (requires -T)
  -1, --fast                 Use fast BAM compression (implies --bam)
  -u, --uncompressed         Uncompressed BAM output (implies --bam)
  -h, --with-header          Include header in SAM output
  -H, --header-only          Print SAM header only (no alignments)
      --no-header            Print SAM alignment records only [default]
  -c, --count                Print only the count of matching records
  -o, --output FILE          Write output to FILE [standard output]
  -U, --unoutput FILE, --output-unselected FILE
                             Output reads not selected by filters to FILE
  -p, --unmap                Set flag to UNMAP on reads not selected
                             then write to output file.
Input options:
  -t, --fai-reference FILE   FILE listing reference names and lengths
  -M, --use-index            Use index and multi-region iterator for regions
      --region[s]-file FILE  Use index to include only reads overlapping FILE
  -X, --customized-index     Expect extra index file argument after 

Filtering options (Only include in output reads that...):
  -L, --target[s]-file FILE  ...overlap (BED) regions in FILE
  -r, --read-group STR       ...are in read group STR
  -R, --read-group-file FILE ...are in a read group listed in FILE
  -N, --qname-file FILE      ...whose read name is listed in FILE
  -d, --tag STR1[:STR2]      ...have a tag STR1 (with associated value STR2)
  -D, --tag-file STR:FILE    ...have a tag STR whose value is listed in FILE
  -q, --min-MQ INT           ...have mapping quality >= INT
  -l, --library STR          ...are in library STR
  -m, --min-qlen INT         ...cover >= INT query bases (as measured via CIGAR)
  -e, --expr STR             ...match the filter expression STR
  -f, --require-flags FLAG   ...have all of the FLAGs present
  -F, --excl[ude]-flags FLAG ...have none of the FLAGs present
  -G FLAG                    EXCLUDE reads with all of the FLAGs present
      --subsample FLOAT      Keep only FLOAT fraction of templates/read pairs
      --subsample-seed INT   Influence WHICH reads are kept in subsampling [0]
  -s INT.FRAC                Same as --subsample 0.FRAC --subsample-seed INT

Processing options:
      --add-flags FLAG       Add FLAGs to reads
      --remove-flags FLAG    Remove FLAGs from reads
  -x, --remove-tag STR
               Comma-separated read tags to strip (repeatable) [null]
      --keep-tag STR
               Comma-separated read tags to preserve (repeatable) [null].
               Equivalent to "-x ^STR"
  -B, --remove-B             Collapse the backward CIGAR operation

General options:
  -?, --help   Print long help, including note about region specification
  -S           Ignored (input format is auto-detected)
      --no-PG  Do not add a PG line
      --input-fmt-option OPT[=VAL]
               Specify a single input file format option in the form
               of OPTION or OPTION=VALUE
  -O, --output-fmt FORMAT[,OPT[=VAL]]...
               Specify output format (SAM, BAM, CRAM)
      --output-fmt-option OPT[=VAL]
               Specify a single output file format option in the form
               of OPTION or OPTION=VALUE
  -T, --reference FILE
               Reference sequence FASTA FILE [null]
  -@, --threads INT
               Number of additional threads to use [0]
      --write-index
               Automatically index the output files [off]
      --verbosity INT
               Set level of verbosity

3.2 格式转换

$ samtools view -@8 -b LPF1_R1_MP.sam > LPF1_R1_MP.bam

-@8：8个线程
-b：输出格式bam文件 >：输出文件名

如果是1.9版本的SAMtools可以参考这篇文章：
https://www.jianshu.com/p/6b7a442d293f
引用转载请注明出处，如有错误敬请指出。