Question: FastQC: 'Per base sequence content' for RNA-Seq
3
gravatar for umn_bist
4.6 years ago by
umn_bist380
umn_bist380 wrote:

Going through my RNA-seq samples, I've noticed this weird pattern for 'per base sequence content' graph.

http://imgur.com/bhYIVJT

There is a lot of noise in the first few base pairs and I was curious what it is and if I should be concerned for not reducing the noise before alignment.

rna-seq • 5.7k views
ADD COMMENTlink modified 4.6 years ago by genomax90k • written 4.6 years ago by umn_bist380
9
gravatar for genomax
4.6 years ago by
genomax90k
United States
genomax90k wrote:

Here is a link (for a post by Dr. Simon Andrews, author of FastQC) to put this topic to rest. Short answer: Nothing to worry about or mitigate.

ADD COMMENTlink modified 4.6 years ago • written 4.6 years ago by genomax90k

Thank you very much!

ADD REPLYlink written 4.6 years ago by umn_bist380

@genomax, so the per base sequence content curves are supposed to be quite close to one another right? What if there is a significant separation between the lines?

ADD REPLYlink written 3 days ago by Dunois140

Can you show an example of what you have?

ADD REPLYlink written 3 days ago by genomax90k

Here you go.

ADD REPLYlink written 3 days ago by Dunois140
1

If your RNA was prepared by poly-A capture/enrichment then an AT bias can exist. What does the GC content plot look like?

ADD REPLYlink modified 3 days ago • written 3 days ago by genomax90k

I think it was poly-A indeed (I still don't know for certain, to be honest). Here's the GC content plot. That one looks fine to me (apart from that bit of a shoulder).

ADD REPLYlink written 3 days ago by Dunois140
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