Dear all,
I am trying to map Illumina 2x126bp exome data (Nimblegen v3) with bowtie2. Subsequently, I want to perform variant detection using Samtools.
I ran into following problem:
- When I do an end-to-end paired-end mapping with bowtie2, the resulting sam file shows a mapping quality (column 5, MAPQ) of 0 for every read.
- When I do a local paired-end mapping with bowtie2, the resulting sam file shows a mapping quality (column 5, MAPQ) of 44 for every read.
- Note: both end-to-end and local mapping of the same reads but as single end reads shows a MAPQ value ranging between 1 and 44 (local) or between 1 and 42 (end-to-end)
Example sam output end-to-end:
bowtie2 --very-sensitive -p 8 --no-mixed --no-discordant -x ./bowtie2_hg19/hg19 -1 F10_GCCAAT_L007_R1.fastq -2 F10_GCCAAT_L007_R2.fastq -S F10_ete.sam --met-file F10_ete.out
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 83 chr19 8841203 0 126M = 8841061 -268 TTAAGTTTTCTCCCTCCAGCCTACTGATTCTAGCTGCAGCCTCATTCCTCTTCTCAAGACACCATCCCAGGAAGCCACCACCAATCAATGATGATTGGCATGTGAGATGAAAATTATTTGCCACCC ####A?9(09.0?88B@A::A>@@>>@A@A>CA::BBBBCCCCA;6(67;7B?7==.)(=)GHFC@=8)<HEBD?700):FHGGFDF?GHGIHHG@IIGF<DH@<H@IGGHF<>FBCABDDDA@@@ AS:i:-4 XN:i:0 XM:i:2 XO:i:0 XG:i:0 NM:i:2 MD:Z:46A31T47 YT:Z:CP
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 163 chr19 8841061 0 126M = 8841203 268 TGAGGCACCACGTCCGGCCGAACCTCAAATTTTTCACTATAGGGAGCAGTGGGTAGCTCTTCATTGGAGTGTGGCCAAGCGTCTCATTCAGAGATGCCAGCTGCTTCTTTGCCATCCCATGCCTTG @<@DFFFDBFBBFBGIIIGIJJJJ<9DH8DFGIIC8BGIEGHII>GIEE7=AA?BDCCCCA@CAA;CA?CDDDD?ACBB?BB?B?@DEECC::A@C4@A@BCC9CCCDDD((4@@A88:@C?::AC AS:i:-2 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:110C15 YT:Z:CP
Example sam output local alignment:
bowtie2 --very-sensitive-local --local -M 5 --score-min G,20,8 -p 8 --no-mixed --no-discordant -x ./bowtie2_hg19/hg19 -1 F10_GCCAAT_L007_R1.fastq -2 F10_GCCAAT_L007_R2.fastq -S F10_local.sam --met-file F10_local.out
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 83 chr19 8841203 44 126M = 8841061 -268 TTAAGTTTTCTCCCTCCAGCCTACTGATTCTAGCTGCAGCCTCATTCCTCTTCTCAAGACACCATCCCAGGAAGCCACCACCAATCAATGATGATTGGCATGTGAGATGAAAATTATTTGCCACCC ####A?9(09.0?88B@A::A>@@>>@A@A>CA::BBBBCCCCA;6(67;7B?7==.)(=)GHFC@=8)<HEBD?700):FHGGFDF?GHGIHHG@IIGF<DH@<H@IGGHF<>FBCABDDDA@@@ AS:i:244 XN:i:0 XM:i:2 XO:i:0 XG:i:0 NM:i:2 MD:Z:46A31T47 YT:Z:CP
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 163 chr19 8841061 44 126M = 8841203 268 TGAGGCACCACGTCCGGCCGAACCTCAAATTTTTCACTATAGGGAGCAGTGGGTAGCTCTTCATTGGAGTGTGGCCAAGCGTCTCATTCAGAGATGCCAGCTGCTTCTTTGCCATCCCATGCCTTG @<@DFFFDBFBBFBGIIIGIJJJJ<9DH8DFGIIC8BGIEGHII>GIEE7=AA?BDCCCCA@CAA;CA?CDDDD?ACBB?BB?B?@DEECC::A@C4@A@BCC9CCCDDD((4@@A88:@C?::AC AS:i:248 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:110C15 YT:Z:CP
Who can make sense of this? Thanks! I already spend a day on this.
Kind regards,
Filip
I am trying to map Illumina 2x126bp exome data (Nimblegen v3) with bowtie2. Subsequently, I want to perform variant detection using Samtools.
I ran into following problem:
- When I do an end-to-end paired-end mapping with bowtie2, the resulting sam file shows a mapping quality (column 5, MAPQ) of 0 for every read.
- When I do a local paired-end mapping with bowtie2, the resulting sam file shows a mapping quality (column 5, MAPQ) of 44 for every read.
- Note: both end-to-end and local mapping of the same reads but as single end reads shows a MAPQ value ranging between 1 and 44 (local) or between 1 and 42 (end-to-end)
Example sam output end-to-end:
bowtie2 --very-sensitive -p 8 --no-mixed --no-discordant -x ./bowtie2_hg19/hg19 -1 F10_GCCAAT_L007_R1.fastq -2 F10_GCCAAT_L007_R2.fastq -S F10_ete.sam --met-file F10_ete.out
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 83 chr19 8841203 0 126M = 8841061 -268 TTAAGTTTTCTCCCTCCAGCCTACTGATTCTAGCTGCAGCCTCATTCCTCTTCTCAAGACACCATCCCAGGAAGCCACCACCAATCAATGATGATTGGCATGTGAGATGAAAATTATTTGCCACCC ####A?9(09.0?88B@A::A>@@>>@A@A>CA::BBBBCCCCA;6(67;7B?7==.)(=)GHFC@=8)<HEBD?700):FHGGFDF?GHGIHHG@IIGF<DH@<H@IGGHF<>FBCABDDDA@@@ AS:i:-4 XN:i:0 XM:i:2 XO:i:0 XG:i:0 NM:i:2 MD:Z:46A31T47 YT:Z:CP
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 163 chr19 8841061 0 126M = 8841203 268 TGAGGCACCACGTCCGGCCGAACCTCAAATTTTTCACTATAGGGAGCAGTGGGTAGCTCTTCATTGGAGTGTGGCCAAGCGTCTCATTCAGAGATGCCAGCTGCTTCTTTGCCATCCCATGCCTTG @<@DFFFDBFBBFBGIIIGIJJJJ<9DH8DFGIIC8BGIEGHII>GIEE7=AA?BDCCCCA@CAA;CA?CDDDD?ACBB?BB?B?@DEECC::A@C4@A@BCC9CCCDDD((4@@A88:@C?::AC AS:i:-2 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:110C15 YT:Z:CP
Example sam output local alignment:
bowtie2 --very-sensitive-local --local -M 5 --score-min G,20,8 -p 8 --no-mixed --no-discordant -x ./bowtie2_hg19/hg19 -1 F10_GCCAAT_L007_R1.fastq -2 F10_GCCAAT_L007_R2.fastq -S F10_local.sam --met-file F10_local.out
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 83 chr19 8841203 44 126M = 8841061 -268 TTAAGTTTTCTCCCTCCAGCCTACTGATTCTAGCTGCAGCCTCATTCCTCTTCTCAAGACACCATCCCAGGAAGCCACCACCAATCAATGATGATTGGCATGTGAGATGAAAATTATTTGCCACCC ####A?9(09.0?88B@A::A>@@>>@A@A>CA::BBBBCCCCA;6(67;7B?7==.)(=)GHFC@=8)<HEBD?700):FHGGFDF?GHGIHHG@IIGF<DH@<H@IGGHF<>FBCABDDDA@@@ AS:i:244 XN:i:0 XM:i:2 XO:i:0 XG:i:0 NM:i:2 MD:Z:46A31T47 YT:Z:CP
HISEQ:126:C0HL2ACXX:7:1101:5336:1952 163 chr19 8841061 44 126M = 8841203 268 TGAGGCACCACGTCCGGCCGAACCTCAAATTTTTCACTATAGGGAGCAGTGGGTAGCTCTTCATTGGAGTGTGGCCAAGCGTCTCATTCAGAGATGCCAGCTGCTTCTTTGCCATCCCATGCCTTG @<@DFFFDBFBBFBGIIIGIJJJJ<9DH8DFGIIC8BGIEGHII>GIEE7=AA?BDCCCCA@CAA;CA?CDDDD?ACBB?BB?B?@DEECC::A@C4@A@BCC9CCCDDD((4@@A88:@C?::AC AS:i:248 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:110C15 YT:Z:CP
Who can make sense of this? Thanks! I already spend a day on this.
Kind regards,
Filip
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