Question: Samtools flagstat results
0
gravatar for banerjeeshayantan
19 months ago by
banerjeeshayantan80 wrote:
221372 + 0 in total (QC-passed reads + QC-failed reads)  
0 + 0 secondary  
0 + 0 supplementary  
20419 + 0 duplicates  
218469 + 0 mapped (98.69% : N/A)  
155851 + 0 paired in sequencing  
77895 + 0 read1  
77956 + 0 read2  
142663 + 0 properly paired (91.54% : N/A)  
150045 + 0 with itself and mate mapped  
2903 + 0 singletons (1.86% : N/A)  
4938 + 0 with mate mapped to a different chr  
2120 + 0 with mate mapped to a different chr (mapQ>=5)

I have the following questions:
1. From previous posts I understood that read1 may not be equal to read2, as there may be reads whose mates didn't align. These are singletons. So doesn't that mean that read2-read1 must be equal to singletons? What am i missing here?
2. What does the field "with itself and mate mapped mean?"

next-gen • 1.1k views
ADD COMMENTlink modified 19 months ago by Devon Ryan88k • written 19 months ago by banerjeeshayantan80

What preprocessing steps have been applied to get this file ?

ADD REPLYlink written 19 months ago by geek_y9.3k

I wrote this command: samtools flagstat example.bam. Does this help?

ADD REPLYlink written 19 months ago by banerjeeshayantan80
1
gravatar for Devon Ryan
19 months ago by
Devon Ryan88k
Freiburg, Germany
Devon Ryan88k wrote:
  1. Singletons occur when only one mate in a pair aligns. You can also have situations where one mate aligns multiple times (e.g., to a simple repeat) and the other only once. Then one will have a single entry and the other may have multiple. Also, if you did any filtering then that'd affect this as well.
  2. It means exactly what is says, both mates mapped. They may be "properly paired" or they may not be. Regardless, if they both align somewhere at least once then they count toward this.
ADD COMMENTlink written 19 months ago by Devon Ryan88k

When I subtract number of reads mapped from the total number of reads (221372-218469=2903), which is the number of singletons. So can I say that singletons are the reads which didn't map to any reference?

ADD REPLYlink written 19 months ago by banerjeeshayantan80

By definition a singleton cannot be unmapped. If it is, it's not a singleton.

ADD REPLYlink written 19 months ago by Devon Ryan88k

Thanks for your reply. Can you explain what you meant by "You can also have situations where one mate aligns multiple times (e.g., to a simple repeat) and the other only once.". I know it is a trivial question, but I am entirely new to this field. Hence asking.

ADD REPLYlink written 19 months ago by banerjeeshayantan80
1

Suppose you have the sequence ATATATATATATATATATAAGCGCTAGCTAGTCGATCTAGCTAGCTGATCGGTCGTCAGAC. You might have reads ATATATAT and GCGCTAGC. The latter read can only align to one place in that sequence. The former read can align equally well to multiple places. Consequently, some aligners will produce multiple entries for ATATATAT and a single one for GCGCTAGC.

ADD REPLYlink written 19 months ago by Devon Ryan88k

Excellent explanation. Thanks a lot!

ADD REPLYlink written 19 months ago by banerjeeshayantan80
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