playing with BWA-MEM : my notebook

BWA-MEM, is a new tool that is part of the latest version of BWA. As said Heng Li on Biostar bwa-mem can be used to identify the viral integration sites in the human genome. Here I've used various short reads to explore how bwa maps the pairs. The sequences I used below are:

• NOTCH2a and NOTCH2b two sequences on the chromosomes 1, on the same strand
• NOTCH2del : same as NOTCH2a but with a deletion
• ROXAN1a and ROXAN1b: two sequences on the chromosome 22, on the same strand
• NOTCH2ins is NOTCH2a with and insertion of ROXAN1a

The following results were generated with the makefile below. Each pair of fastq was submitted twice (1_* and 2_*). I also added some pairs of reads from another source of FASTQs to let bwa infer the size of the fragments (see biostar: http://www.biostars.org/p/69694).

NOTCH2a vs NOTCH2b BWA-MEM options: none

NOTCH2a vs NOTCH2b
BWA-MEM options: none
NOT Properly mapped reads because they're on the same strand.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2	p1=65	1	120572000	9	50M	NM:i:0 AS:i:50 XS:i:45
1_Notch2	p2=129	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2	p1	1	120572000	9	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2	p2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2a vs revcomp(NOTCH2b) BWA-MEM options: none

NOTCH2a vs revcomp(NOTCH2b)
BWA-MEM options: none
Properly mapped reads.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2	pPR1=99	1	120572000	39	50M	NM:i:0 AS:i:50 XS:i:45
1_Notch2	pPr2=147	1	120572200	39	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2	pPR1	1	120572000	39	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2	pPr2	1	120572200	39	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2del vs revcomp(NOTCH2b) BWA-MEM options: none

NOTCH2del vs revcomp(NOTCH2b)
BWA-MEM options: none
Deletion in Read1

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2del	pPR1=99	1	120572000	39	37M20D31M	NM:i:20 AS:i:42 XS:i:37
1_Notch2del	pPr2=147	1	120572200	39	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2del	pPR1	1	120572000	39	37M20D31M	NM:i:20 AS:i:42 XS:i:37
2_Notch2del	pPr2	1	120572200	39	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2ins vs revcomp(NOTCH2b) BWA-MEM options:

NOTCH2ins vs revcomp(NOTCH2b)
BWA-MEM options: non
Soft clipping of chr1: two hits on different chromosomes.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_NOTCH2insRoxan	pR1=97	22	41721566	60	23S50M27S	NM:i:0 AS:i:50 XS:i:0
1_NOTCH2insRoxan	pr2=145	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45
2_NOTCH2insRoxan	pR1	22	41721566	60	23S50M27S	NM:i:0 AS:i:50 XS:i:0
2_NOTCH2insRoxan	pr2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b) BWA-MEM options: none

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b)
BWA-MEM options: none
two hits on chromosome chr1 and chr22 for the 5' seq
nevertheless, properly paired was set.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2Roxan	pPR1=99	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
1_Notch2Roxan	pPR1=99	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
1_Notch2Roxan	pPr2=147	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2Roxan	pPR1	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
2_Notch2Roxan	pPR1	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
2_Notch2Roxan	pPr2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b+ROXAN1b) BWA-MEM options: none

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b+ROXAN1b)
BWA-MEM options: none
All fragments on chr1/chr22.
Reads are NOT properly paired.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2Roxan	pR1=97	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
1_Notch2Roxan	pR1=97	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
1_Notch2Roxan	pr2=145	22	41721766	60	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,-120572200,50M50S,9,0;
1_Notch2Roxan	pr2=145	1	120572200	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,-41721766,50S50M,60,0;
2_Notch2Roxan	pR1	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
2_Notch2Roxan	pR1	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
2_Notch2Roxan	pr2	22	41721766	60	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,-120572200,50M50S,9,0;
2_Notch2Roxan	pr2	1	120572200	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,-41721766,50S50M,60,0;

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b) BWA-MEM options: -C

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b)
BWA-MEM options: -C
I cannot see the effect of the option '-C':
"append FASTA/FASTQ comment to SAM output"

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2Roxan	pPR1=99	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
1_Notch2Roxan	pPR1=99	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
1_Notch2Roxan	pPr2=147	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2Roxan	pPR1	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
2_Notch2Roxan	pPR1	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
2_Notch2Roxan	pPr2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b) BWA-MEM options: -M

NOTCH2a+ROXAN1a vs revcomp(NOTCH2b)
BWA-MEM options: -M
Set the duplicate flags for the multiple hits.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_Notch2Roxan	pPR1=99	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
1_Notch2Roxan	pPR1s=355	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
1_Notch2Roxan	pPr2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45
2_Notch2Roxan	pPR1	1	120572000	9	50M50S	NM:i:0 AS:i:50 XS:i:45 XP:Z:22,+41721566,50S50M,9,0;
2_Notch2Roxan	pPR1s	22	41721566	9	50S50M	NM:i:0 AS:i:50 XS:i:0 XP:Z:1,+120572000,50M50S,9,0;
2_Notch2Roxan	pPr2	1	120572200	9	50M	NM:i:0 AS:i:50 XS:i:45

NOTCH2ins vs revcomp(ROXAN1b) BWA-MEM options: none

NOTCH2ins vs revcomp(ROXAN1b)
BWA-MEM options: none
Reads are mapped on chr22.

Read	Flag	Chrom	Pos	qual	Cigar	Extra
1_NOTCH2insRoxan	pPR1	22	41721566	60	23S50M27S	NM:i:0 AS:i:50 XS:i:0
1_NOTCH2insRoxan	pPr2	22	41721766	60	50M	NM:i:0 AS:i:50 XS:i:0
2_NOTCH2insRoxan	pPR1	22	41721566	60	23S50M27S	NM:i:0 AS:i:50 XS:i:0
2_NOTCH2insRoxan	pPr2	22	41721766	60	50M	NM:i:0 AS:i:50 XS:i:0

That's it,

Pierre

The makefile