SAMtools的常用指令


1.Samtools的help:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools --help

Program: samtools (Tools for alignments in the SAM format)
Version: 1.3-20-gd49c73b (using htslib 1.3-35-g26b3085)

Usage:   samtools <command> [options]

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

  -- Editing
     calmd          recalculate MD/NM tags and '=' bases
     fixmate        fix mate information
     reheader       replace BAM header
     rmdup          remove PCR duplicates
     targetcut      cut fosmid regions (for fosmid pool only)
     addreplacerg   adds or replaces RG tags

  -- 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

  -- Statistics
     bedcov         read depth per BED region
     depth          compute the depth
     flagstat       simple stats
     idxstats       BAM index stats
     phase          phase heterozygotes
     stats          generate stats (former bamcheck)

  -- Viewing
     flags          explain BAM flags
     tview          text alignment viewer
     view           SAM<->BAM<->CRAM conversion
     depad          convert padded BAM to unpadded BAM


2.Bam和Sam相互转换:

 samtools view -h file.bam > file.sam
 samtools view -b -S file.sam > file.bam

示例:

samtools view -h NA12878.bam >NA12878_2.sam
samtools view -b -S NA12878.sam > NA12878_2.bam


3.构建索引index:

samtools index sorted.bam
实例:

samtools index NA12878.bam
生成NA12878.bam.bai,具体:

不知道如何查看.bai文件???

4.统计信息flagstat:

 samtools flagstat NA12878Merge.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools flagstat NA12878Merge.bam 
1058 + 72 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
0 + 0 supplementary
118 + 32 duplicates
1058 + 72 mapped (100.00% : 100.00%)
1058 + 72 paired in sequencing
516 + 26 read1
542 + 46 read2
1048 + 64 properly paired (99.05% : 88.89%)
1048 + 64 with itself and mate mapped
10 + 8 singletons (0.95% : 11.11%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)



大文件:

hadoop@Mcnode6:~/cloud/adam/xubo/1000genomes/GIH/NA21144/alignment$ samtools flagstat NA21144.alt_bwamem_GRCh38DH.20150718.GIH.low_coverage.cram
247384356 + 0 in total (QC-passed reads + QC-failed reads)
2090839 + 0 secondary
9082475 + 0 supplementary
5797777 + 0 duplicates
246212003 + 0 mapped (99.53% : N/A)
236211042 + 0 paired in sequencing
118105521 + 0 read1
118105521 + 0 read2
226096416 + 0 properly paired (95.72% : N/A)
233890591 + 0 with itself and mate mapped
1148098 + 0 singletons (0.49% : N/A)
4379358 + 0 with mate mapped to a different chr
1561618 + 0 with mate mapped to a different chr (mapQ>=5)




5.合并文件merge:

bam文件:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools merge NA12878Merge.bam NA12878.bam NA12878_2.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ls
aln.sorted.bam  cramtools-3.0.jar  NA12878_2.bam  NA12878_2.sam  NA12878.bam  NA12878.bam.bai  NA12878Merge.bam  NA12878.sam
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ll
total 4676
drwxrwxr-x  2 hadoop hadoop    4096  3月  9 20:41 ./
drwx------ 13 hadoop hadoop    4096  3月  7 22:07 ../
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 20:29 aln.sorted.bam
-rw-rw-r--  1 hadoop hadoop 3978747  3月  8 22:01 cramtools-3.0.jar
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 20:27 NA12878_2.bam
-rw-rw-r--  1 hadoop hadoop  255818  3月  9 16:06 NA12878_2.sam
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 16:00 NA12878.bam
-rw-rw-r--  1 hadoop hadoop     200  3月  9 16:04 NA12878.bam.bai
-rw-r--r--  1 hadoop hadoop    4096  3月  9 16:01 .NA12878.bam.swp
-rw-rw-r--  1 hadoop hadoop   75781  3月  9 20:41 NA12878Merge.bam
-rw-r--r--  1 hadoop hadoop  255818 11月 16  2014 NA12878.sam

bam统计信息:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools flagstat NA12878.bam
529 + 36 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
0 + 0 supplementary
59 + 16 duplicates
529 + 36 mapped (100.00% : 100.00%)
529 + 36 paired in sequencing
258 + 13 read1
271 + 23 read2
524 + 32 properly paired (99.05% : 88.89%)
524 + 32 with itself and mate mapped
5 + 4 singletons (0.95% : 11.11%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools flagstat NA12878Merge.bam
1058 + 72 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
0 + 0 supplementary
118 + 32 duplicates
1058 + 72 mapped (100.00% : 100.00%)
1058 + 72 paired in sequencing
516 + 26 read1
542 + 46 read2
1048 + 64 properly paired (99.05% : 88.89%)
1048 + 64 with itself and mate mapped
10 + 8 singletons (0.95% : 11.11%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)


sam文件:
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools merge NA12878Merge.sam NA12878.sam NA12878_2.sam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ls
aln.sorted.bam  cramtools-3.0.jar  NA12878_2.bam  NA12878_2.sam  NA12878.bam  NA12878.bam.bai  NA12878Merge.bam  NA12878Merge.sam  NA12878.sam
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ll
total 5188
drwxrwxr-x  2 hadoop hadoop    4096  3月  9 20:44 ./
drwx------ 13 hadoop hadoop    4096  3月  7 22:07 ../
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 20:29 aln.sorted.bam
-rw-rw-r--  1 hadoop hadoop 3978747  3月  8 22:01 cramtools-3.0.jar
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 20:27 NA12878_2.bam
-rw-rw-r--  1 hadoop hadoop  255818  3月  9 16:06 NA12878_2.sam
-rw-rw-r--  1 hadoop hadoop   64321  3月  9 16:00 NA12878.bam
-rw-rw-r--  1 hadoop hadoop     200  3月  9 16:04 NA12878.bam.bai
-rw-r--r--  1 hadoop hadoop    4096  3月  9 16:01 .NA12878.bam.swp
-rw-rw-r--  1 hadoop hadoop   75781  3月  9 20:41 NA12878Merge.bam
-rw-rw-r--  1 hadoop hadoop  521677  3月  9 20:44 NA12878Merge.sam
-rw-r--r--  1 hadoop hadoop  255818 11月 16  2014 NA12878.sam

sam文件统计信息:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools flagstat NA12878Merge.sam 
1058 + 72 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
0 + 0 supplementary
118 + 32 duplicates
1058 + 72 mapped (100.00% : 100.00%)
1058 + 72 paired in sequencing
516 + 26 read1
542 + 46 read2
1048 + 64 properly paired (99.05% : 88.89%)
1048 + 64 with itself and mate mapped
10 + 8 singletons (0.95% : 11.11%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools flagstat NA12878.sam 
529 + 36 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
0 + 0 supplementary
59 + 16 duplicates
529 + 36 mapped (100.00% : 100.00%)
529 + 36 paired in sequencing
258 + 13 read1
271 + 23 read2
524 + 32 properly paired (99.05% : 88.89%)
524 + 32 with itself and mate mapped
5 + 4 singletons (0.95% : 11.11%)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)

6. 排序sort:

samtools sort -o NA12878.Sorted.bam NA12878.bam


7.索引统计idxstats:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools index aln.sorted.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools idxstats aln.sorted.bam
20	63025520	565	0
*	0	0	0
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ls
aln.sorted.bam  aln.sorted.bam.bai  cramtools-3.0.jar  NA12878_2.bam  NA12878_2.sam  NA12878.bam  NA12878.bam.bai  NA12878Merge.bam  NA12878Merge.sam  NA12878.sam  NA12878.Sorted.bam

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools view -b -S small.sam >small.bam
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ ls
artificial.fa             fastq_sample1.fq                       interleaved_fastq_sample1.ifq         proper_pairs_2.fq                    small_realignment_targets.pileup
artificial.README.txt     fastq_sample2.fq                       interleaved_fastq_sample1.ifq.output  reads-0-2-0                          small_realignment_targets_README.txt
artificial.realigned.sam  fastq_sample3.fq                       interleaved_fastq_sample2.ifq         reads12_diff1.sam                    small_realignment_targets.sam
artificial.sam            fastq_sample4.fq                       interleaved_fastq_sample2.ifq.output  reads12.sam                          small.sam
bqsr1-ref.observed        features                               interleaved_fastq_sample3.ifq         reads21.sam                          small.vcf
bqsr1.sam                 gencode.v19.pc_transcripts.250k.fa.gz  interleaved_fastq_sample3.ifq.output  single_fastq_sample1.fq.output       sorted.sam
bqsr1.snps                hg19.chrM.2bit                         interleaved_fastq_sample4.ifq         single_fastq_sample2.fq.output       tags.sam
bqsr1.vcf                 Hs_Ensembl_example_genes.gtf           interleaved_fastq_sample4.ifq.output  single_fastq_sample3.fq.output       test.conf
chr20.250k.fa.gz          human_g1k_v37_chr1_59kb.2bit           multiline_fastq.fq                    single_fastq_sample4.fq.output       test_rowgroup_rangeindex.1.txt
dict_with_accession.dict  human_g1k_v37_chr1_59kb.fasta          ordered.sam                           small.bam                            unmapped.sam
example_intervals.list    improper_pairs_1.fq                    parquet_lister_dir_empty              small_missing.vcf                    unordered.sam
fastq_noqual.fq           improper_pairs_2.fq                    proper_pairs_1.fq                     small_realignment_targets.intervals  unsorted.sam
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools sort -o smallSort.bam small.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools index smallSort.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools idxstats smallSort.bam
1	249250621	20	0
2	243199373	0	0
*	0	0	0


7.深度depth:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools depth --help
depth: unrecognized option '--help'

Usage: samtools depth [options] in1.bam [in2.bam [...]]
Options:
   -a                  output all positions (including zero depth)
   -a -a (or -aa)      output absolutely all positions, including unused ref. sequences
   -b <bed>            list of positions or regions
   -f <list>           list of input BAM filenames, one per line [null]
   -l <int>            read length threshold (ignore reads shorter than <int>)
   -d/-m <int>         maximum coverage depth [8000]
   -q <int>            base quality threshold
   -Q <int>            mapping quality threshold
   -r <chr:from-to>    region
      --input-fmt-option OPT[=VAL]
               Specify a single input file format option in the form
               of OPTION or OPTION=VALUE
      --reference FILE
               Reference sequence FASTA FILE [null]

The output is a simple tab-separated table with three columns: reference name,
position, and coverage depth.  Note that positions with zero coverage may be
omitted by default; see the -a option.

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam/resources$ samtools depth small.bam 
1	26472784	1
1	26472785	1
1	26472786	1
1	26472787	1
1	26472788	1
1	26472789	1
1	26472790	1
1	26472791	1
1	26472792	1
1	26472793	1
1	26472794	1
1	26472795	1
1	26472796	1
1	26472797	1
1	26472798	1
1	26472799	1
1	26472800	1
1	26472801	1
1	26472802	1
1	26472803	1
1	26472804	1
1	26472805	1
1	26472806	1
1	26472807	1
1	26472808	1
1	26472809	1
1	26472810	1
1	26472811	1
1	26472812	1
1	26472813	1
1	26472814	1
1	26472815	1
1	26472816	1
1	26472817	1
1	26472818	1
1	26472819	1
1	26472820	1
1	26472821	1
1	26472822	1
1	26472823	1
1	26472824	1
1	26472825	1
1	26472826	1
1	26472827	1
1	26472828	1
1	26472829	1
1	26472830	1
1	26472831	1
1	26472832	1
1	26472833	1
1	26472834	1
1	26472835	1
1	26472836	1
1	26472837	1
1	26472838	1
1	26472839	1
1	26472840	1
1	26472841	1
1	26472842	1
1	26472843	1
1	26472844	1
1	26472845	1
1	26472846	1
1	26472847	1
1	26472848	1
1	26472849	1
1	26472850	1
1	26472851	1
1	26472852	1
1	26472853	1
1	26472854	1
1	26472855	1
1	26472856	1
1	26472857	1
1	26472858	1

8.给fa文件建立索引faidx:

faidx

    samtools faidx <ref.fasta> [region1 [...]]

    Index reference sequence in the FASTA format or extract subsequence 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 specified, the subsequences will be retrieved and printed to stdout in the FASTA format.

    The input file can be compressed in the BGZF format.

    The sequences in the input file should all have different names. If they do not, indexing will emit a warning about duplicate sequences and retrieval will only produce subsequences from the first sequence with the duplicated name. 

hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ ls
00README.txt  ex1.fa  ex1.sam.gz  toy.fa  toy.sam
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools faidx ex1.fa 
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ ls
00README.txt  ex1.fa  ex1.fa.fai  ex1.sam.gz  toy.fa  toy.sam


9.text查看比对图tview:

 samtools tview [-p chr:pos] [-s STR] [-d display] <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 alignment start from a region in the format like `chr10:10,000,000' or `=10,000,000' when viewing the same reference sequence.

Options:

-d display

    Output as (H)tml or (C)urses or (T)ext 
-p chr:pos

    Go directly to this position 
-s STR

    Display only alignments from this sample or read group 

运行:

hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools view -h -S toy.sam >toy.bam
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools sort -o toy.sorted.bam toy.bam
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools tview  toy.bam toy.fa
Cannot read index for 'toy.bam'.
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools index toy.sorted.bam 
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ ls
00README.txt  ex1.fa  ex1.fa.fai  ex1.sam.gz  toy.bam  toy.fa  toy.fa.fai  toy.sam  toy.sorted.bam  toy.sorted.bam.bai  toy.sorted.sam
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools tview  toy.sbam toy.fa
toy.sam             toy.sorted.bam      toy.sorted.bam.bai  toy.sorted.sam      
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools tview  toy.sbam toy.fa
toy.sam             toy.sorted.bam      toy.sorted.bam.bai  toy.sorted.sam      
hadoop@Mcnode1:~/cloud/adam/xubo/code/samtools/examples$ samtools tview  toy.sorted.bam toy.fa

1         11              21        31         41        51        61        71        81        91        101       111       121       131       141       151       161       171
AGCATGTTAGATAA****GATA**GCTGTGCTAGTAGGCAG*TCAGCGCCATNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN
      ........    ....  ......K.K......K. ..........
      ........AGAG....***...      ,,,,,    ,,,,,,,,,
        ......GG**....AA
        ..C...**** ...**...>>>>>>>>>>>>>>T.....


10.将bam/sam/cram文件转换成fasta/fastq文件:

hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools fasta -s NA12878.fa  NA12878.sam 
[M::bam2fq_mainloop_singletontrack] discarded 565 singletons
[M::bam2fq_mainloop_singletontrack] processed 565 reads
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ ls
aln.sorted.bam      cramtools-3.0.jar  NA12878_2.sam  NA12878.bam.bai  NA12878Merge.bam  NA12878.sam         resources
aln.sorted.bam.bai  NA12878_2.bam      NA12878.bam    NA12878.fa       NA12878Merge.sam  NA12878.Sorted.bam
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ vi NA12878.fa
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ samtools fastq -s NA12878.fq  NA12878.sam 
[M::bam2fq_mainloop_singletontrack] discarded 565 singletons
[M::bam2fq_mainloop_singletontrack] processed 565 reads
hadoop@Mcnode1:~/cloud/adam/xubo/data/data_HDFS/adam$ more NA12878.fq
@20FUKAAXX100202:3:6:15018:84106/2
ACCCAAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAA
+
?@BBBCEEDFEFEEEFDEEFEEEEBFEDEFCFDDEEFEDFDFEEEFEEEECEEFEEFCEFDEEFFEFEDEEEFFFDECEDCEFEEDDFFBFEFGEAEDCCC
@20GAVAAXX100126:8:62:5578:2527/1
TTGCTGATTGAATCCCCATGGTGGTATTTCATGATCTTCCTTTGTCCTTGGTATATCCTAGAGCTTGGGAGTTAGAGTGGGAGCCTTGATTAGCTTTGGGT
+
1;@3@?<>>?<;:=;==?=>9=?9>9===>=<<;;@@BAABAA>B>4?><8<:=<==;>9:@?>=;<7+/;:/80?5>;-(-;89<::8::8:(1,/:438
@20FUKAAXX100202:4:47:20584:49257/2
CCAAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAAT
+
?ACDBBCEDFEDEFEEEFEDBECFBFEFCFDEEEFEDFDFEEEFEEEECEEFEEFCEFFEEFFEFEDEAEFFFAECEFCDFEEFBFFDBEEC:@6A?C4>B
@20GAVAAXX100126:7:47:4730:37293/2
CCAAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAAT
+
?BB@BCBFDDECC=E@@DB;BDCFDE<<AEB@B>BADD>?C?EDEB>@AC=<?=DAE?E=CAC?;<C=@ADD?ACACCAC>:>4=B676<17@@<:AA<;6
@20GAVAAXX100126:5:46:21151:39489/1
ATTTGCTGATTGAATCCCCATGGTGGTATTTCATGATCTTCCTTTGTCCTTGGTATATCCTAGAGCTTGGGAGTTAGAGTGGGAGCCTTGATTAGATTTGG
+
A=DED=FEFEFEFFEEEEEFEEEEEEEEEFFDEEDFEDFFDDFFFDDDDFFEEDEEEEDDFEEFEEFFEEEFEDFEEFECEEEFEEEFFE>BB>BBB=<9>
@20FUKAAXX100202:6:2:2264:85470/1
CAAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAATT
+
ABDFEEFEGEEFFEFEFEEEFCCDFEFCFEEEEFEDFDF?EEFCEEEBCEFECFCEFFEDFFBE=CF=CFEFDEB@D=CDDC>D:?;;=@DACECAD5A>9
@20FUKAAXX100202:7:24:21050:192748/1
CAAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAATT
+
ABEFEEFFGFEFFEEEFEEEFCFDFEFCFEEEEFEDFDFEEEFEEEECDEFEEFCEFFEEFFEFCCFBE@FGDEAAEBCDC@:FBFBB?EDEEDCCDCB=<
@20FUKAAXX100202:6:48:15418:77607/1
AAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAATTC
+
ACDEDFEGEEFGEEEFEEEECFEFEFCFEEEEFEDFDFEEEFEEEECEEFEEFCEFFEEFFEFEDEEEFFFEECEFCEFFEFFFDDFEFGFEFFEEFECCB
@20GAVAAXX100126:3:65:13772:1294/1
AAATCTAATCAAGGCTCCCACTCTAACTCCCAAGCTCTAGGATATACCAAGGACAAAGGAAGATCATGAAATACCACCATGGGGATTCAATCAGCAAATTC
+



详细操作请参考【1】





参考资料:

【1】 http://www.bbioo.com/lifesciences/40-113338-1.html

【2】 http://www.htslib.org/doc/samtools.html

【3】

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