N reads

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Wafaa al-Aamri

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Sep 22, 2021, 1:43:31 PM9/22/21
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Hello,

I am analyzing my samples on IGV and more than half of them are read as "N". example below

A:0

C:0

G:72 (26%)

T: 0

N: 204

Any idea why is this happening and I also realized IGV doesn't call the variant?

Thank you

James Robinson

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Sep 23, 2021, 10:45:42 PM9/23/21
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I do not understand why more than 1/2 of your read pileup at that location has "N" for the readbase, but IGV is just reporting what it finds in the bam file.   If you think that is not the case we will need a sample of your bam file,  you can use samtools for that or IGV itself (use the "export alignments" option in the alignment right-click menu). 

By default IGV will only call a variant if > 20% of quality-weighted bases mismatches the reference.   26% is a gross total, if some of those are of low quality this could be the reason.  You can change this threshold in the alignment preferences.

Wafaa al-Aamri

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Sep 24, 2021, 2:53:20 PM9/24/21
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Hello James,

Thank you for your reply. I changed the mapping quality threshold. It was set to "0". I changed it to "20". 
Now, my "N" is 0 , and the coverage dropped for most samples. 
I tried to attach the bam file, but it says its too big to be sent. Is there an alternative way (or email) i can send the bam file. Thank you so much.

Wafa



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James Robinson

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Sep 24, 2021, 10:26:07 PM9/24/21
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Hi,  what exactly is your question?  

On Fri, Sep 24, 2021 at 3:40 PM Wafaa al-Aamri <w.al...@gmail.com> wrote:
Hello James,

I think I was able to compress the files, please have a look at our bam file, Thank you so much

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mousumee khan

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Sep 27, 2021, 1:04:41 AM9/27/21
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Hello,

Can anyone tell me how to find out ambiguous sequence (like W, S, Y, K, V, H, D, B, M, N, R etc..) areas in IGV software? Thanks in advance. 

robin...@gmail.com

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Sep 28, 2021, 11:07:36 AM9/28/21
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Hi,

When posting a new question its best fo start a new topic, otherwise the conversation thread gets very confusing to follow.   

I'm not sure I understand your question, what is it exactly that you want to do?   

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