Question: Unexplained Unmapped Read Mates Produced By Tophat2
3
gravatar for mathieu.bahin
5.2 years ago by
France
mathieu.bahin40 wrote:

Hi all,

I have a SAM file produced by TopHat 2 (version 2.0.8) in which I find these two following mates that I can't explain:

HISEQ4_0101:8:1101:10030:81268#TGACCA   73      17      22696364        50      67M     *       0       0       ATGGATATGAATTTTTTGCTAAACGACAATTGGTAACCCTATTTTCAGCCCCAAATTACTGTGGCGA  CC@FFFFFHHHHHJJIJJJJJJJJIDGEIEIJDBDGICHCGIJJIJIJIJJIHJJIGHIEIHEFCD?     MD:Z:67 RG:Z:1  XG:i:0  NH:i:1  NM:i:0  XM:i:0  XN:i:0  XO:i:0       AS:i:0  XS:A:+  YT:Z:UU

HISEQ4_0101:8:1101:10030:81268#TGACCA   153     17      22696364        50      101M    *       0       0       ATGGATATGAATTTTTTGCTAAACGACAATTGGTAACCCTATTTTCAGCCCCAAATTACTGTGGCGAGTTTGATAATGCTGGTGGAATGATGAGTGTGGAT        DDEDDEEDDDBDBDDDDDDDDDDDEEEFFEFFFHE?FIHGGGHGJJIJJIIHIJGHDBJJJIGHHHCIIIIIJIJIIIJJJIJJJJJIHHGHGFFFFDBB<        MD:Z:101        RG:Z:1  XG:i:0  NH:i:1  NM:i:0  XM:i:0  XN:i:0  XO:i:0  AS:i:0  XS:A:+  YT:Z:UU
  • Same read ID
  • the flags show a first (73) and a second (153) in pair
  • Mapped on the same chromosome
  • Mate unmapped (7th field = "*")
  • NH:i:1 for both (no multiple alignment)

Why is there a star in the 7th field instead of an equal as expected ?

Cheers

tophat2 sam bam • 2.1k views
ADD COMMENTlink modified 5.2 years ago by Pierre Lindenbaum116k • written 5.2 years ago by mathieu.bahin40

You might check what settings tophat2 is using when it's running bowtie2. That might be considered a "dovetail" mapping in bowtie2, so it wouldn't be valid for mates by default (try feeding that to bowtie2 by itself while playing with the settings to check this).

ADD REPLYlink written 5.2 years ago by Devon Ryan88k
0
gravatar for Pierre Lindenbaum
5.2 years ago by
France/Nantes/Institut du Thorax - INSERM UMR1087
Pierre Lindenbaum116k wrote:

there is a big problem with tophat2:

And I'm still waiting for an answer...

ADD COMMENTlink written 5.2 years ago by Pierre Lindenbaum116k

Did you ever try feeding a few of those reads (only non-spliced ones) into bowtie2 while playing with the --dovetail option? I suspect that it's bowtie2 that's actually the issue here.

ADD REPLYlink written 5.2 years ago by Devon Ryan88k

I tried to align the two reads with bowtie2 only (+default opts). The SAM flags were OK.

ADD REPLYlink written 5.2 years ago by Pierre Lindenbaum116k
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