Question: Unaligned Reads From Bowtie2
1
gravatar for deepthithomaskannan
5.3 years ago by
Canada
deepthithomaskannan250 wrote:

Hi all , I am using bowtie2 for mapping illumina paired end mapping. Is there any option for getting the reads which are aligned neither concordantly nor dicordantly? All my reads have pairs. From the manual I infered that --un-conc can be used to get the paired end reads which are failed to align concordantly.

In paired end read alignment whether we have to completely ignore the discordant reads? How much importance we have to give the discordant reads?

Following is the sample of my bowtie2 status summary:

465909 reads; of these:

465909 (100.00%) were paired; of these:

61524 (13.21%) aligned concordantly 0 times
322977 (69.32%) aligned concordantly exactly 1 time
81408 (17.47%) aligned concordantly >1 times
----
61524 pairs aligned concordantly 0 times; of these:
  9488 (15.42%) aligned discordantly 1 time
----
52036 pairs aligned 0 times concordantly or discordantly; of these:
  104072 mates make up the pairs; of these:
    67801 (65.15%) aligned 0 times
    16898 (16.24%) aligned exactly 1 time         (Question:If these pairs are aligned 0 times concordantly or discordantly, then in what way they are aligned? )
    19373 (18.61%) aligned >1 times

92.72% overall alignment rate

Thanks, D

bowtie2 • 4.0k views
ADD COMMENTlink written 5.3 years ago by deepthithomaskannan250

Hi, did you check this --un option instead of --un-conc

ADD REPLYlink written 5.3 years ago by Prakki Rama2.2k

--un options will output ulaigned unpaired sequences. In this case I am getting nothing. Because all of my reads are paired. I have to get unaligned reads eventhough they are paired.

ADD REPLYlink written 5.3 years ago by deepthithomaskannan250

I think the output file will have all the unaligned reads (both paired and single). You need to separate the all the read1's into 1 file and read2's into other file and the remaining will be unaligned reads with only one of the pair hitting to reference.

ADD REPLYlink modified 5.3 years ago • written 5.3 years ago by Prakki Rama2.2k

Reading the bowtie2 manual, "These reads correspond to the SAM records with the FLAGS 0x4 bit set and neither the0x40 nor 0x80 bits set." This means that the --un option outputs only unpaired reads, even though it searches for unpaired alignments when reads fail to align either concordantly or discordantly. I am having the same issue deepthithomaskannan

ADD REPLYlink written 4.5 years ago by mrals8910

Anybody have any idea about the third set in the summary. How the 52036 pairs are aligned? They said they are neither aligned concordantly nor discordantly.

ADD REPLYlink modified 4.5 years ago • written 4.5 years ago by deepthithomaskannan250

I think I got it now.  sometimes only one of the read in the pair has a unique alignment.
 

ADD REPLYlink written 4.5 years ago by deepthithomaskannan250

I assume the --un option will give you the 67801 reads which are unaligned.

ADD REPLYlink written 4.5 years ago by deepthithomaskannan250
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