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HISAT2 alignment, read 2 always is aligning lower than read 1 as exactly 1 time alignment

Dear all,

I have been aligning my paired end sequenced data to the reference genome using HISAT2. I noticed the that read 2 is always less aligned as exactly one time to the reference genome. is it something normal or I should consider some other details while I am aligning.

Thank you

rna-seq
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updated by Ram Ram • written by e.amiri79 • e.amiri79

read 2 is always less aligned as exactly one time to the reference genome

I do not understand what you mean with this. It is normal that read 2 is of lower quality though.

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written by WouterDeCoster WouterDeCoster

Do you think it is a normal result I got after hisat2 alignment?

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written by e.amiri79 • e.amiri79

You need to show us the numbers you're talking about.

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written by Devon Ryan Devon Ryan 
`enter code here`13837382 reads; of these:
  13837382 (100.00%) were paired; of these:
    4356417 (31.48%) aligned concordantly 0 times
    9330004 (67.43%) aligned concordantly exactly 1 time
    150961 (1.09%) aligned concordantly >1 times
    ----
    4356417 pairs aligned concordantly 0 times; of these:
      70630 (1.62%) aligned discordantly 1 time
    ----
    4285787 pairs aligned 0 times concordantly or discordantly; of these:
      8571574 mates make up the pairs; of these:
        7100692 (82.84%) aligned 0 times
        1419865 (16.56%) aligned exactly 1 time
        51017 (0.60%) aligned >1 times
74.34% overall alignment rate`enter code here`
• 0 views • link
written by e.amiri79 • e.amiri79

There's nothing about read 2 there

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written by Devon Ryan Devon Ryan

If you notice the alignment summery, the read 2 aligned almost half of the read one to the reference genome

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written by e.amiri79 • e.amiri79

Please use ADD COMMENT/ADD REPLY when responding to existing posts to keep threads logically organized.

BTW: Where is the alignment summary? Nothing appears to be included in this thread in terms of stats/images.

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written by GenoMax GenoMax

If you're getting this from samtools flagstat or something like that then please post those metrics.

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written by Devon Ryan Devon Ryan 
13837382 reads; of these:
  13837382 (100.00%) were paired; of these:
    4356417 (31.48%) aligned concordantly 0 times
    9330004 (67.43%) aligned concordantly exactly 1 time
    150961 (1.09%) aligned concordantly >1 times
    ----
    4356417 pairs aligned concordantly 0 times; of these:
      70630 (1.62%) aligned discordantly 1 time
    ----
    4285787 pairs aligned 0 times concordantly or discordantly; of these:
      8571574 mates make up the pairs; of these:
        7100692 (82.84%) aligned 0 times
        1419865 (16.56%) aligned exactly 1 time
        51017 (0.60%) aligned >1 times
74.34% overall alignment rate
• 0 views • link
updated by WouterDeCoster WouterDeCoster • written by e.amiri79 • e.amiri79

AGAIN

Please use ADD COMMENT or ADD REPLY to answer to previous reactions, as such this thread remains logically structured and easy to follow. I have now moved your reaction but as you can see it's not optimal. Adding an answer should only be used for providing a solution to the question asked.

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written by WouterDeCoster WouterDeCoster

I added code markup to your post for increased readability. You can do this by selecting the text and clicking the 101010 button. When you compose or edit a post that button is in your toolbar, see image below:

101010 Button

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written by WouterDeCoster WouterDeCoster

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