SEQanswers

Go Back   SEQanswers > General



Similar Threads
Thread Thread Starter Forum Replies Last Post
CDS vs cDNA s13ep Complete Genomics 4 04-20-2012 02:19 PM
Structural variation and implications for PacBio RS? ETHANol Literature Watch 0 08-09-2011 12:47 AM
Exon-Junction mapping: re-assigning CDS-mapped reads to chromosomes sridharacharya RNA Sequencing 1 10-21-2010 04:07 PM
comparing sequences and mapping annotations? gaster Bioinformatics 0 07-06-2009 01:33 PM

Reply
 
Thread Tools
Old 06-30-2011, 12:58 AM   #1
sisch
Member
 
Location: Dusseldorf, Germany

Join Date: Jun 2011
Posts: 29
Default [solved] Implications from comparing cDNA to cds mapping

Hey there,

I've had an RNA Seq experiment on the 454 platform and during data analysis I came to wonder:

I mapped my reads (non-model organism) to a closely related genome using cDNA and coding sequences(cds). The fraction of mappable reads is 0.4 in cds and 0.6 in cDNA. So now I'm wondering whether this does infer that the 20% reads that mapped to cdna but not to cds are non-mRNA fragments (contamination of some sorts) or whether this implication is wrong

I feel like I'm overlooking something when I draw this conclusion but I can't point the finger to it.

Best,
Simon

Last edited by sisch; 07-13-2011 at 07:50 AM. Reason: adding 'solved'-tag
sisch is offline   Reply With Quote
Old 06-30-2011, 08:09 AM   #2
husamia
Member
 
Location: cinci

Join Date: Apr 2010
Posts: 66
Default

what you mean by cDNA?
husamia is offline   Reply With Quote
Old 06-30-2011, 08:25 AM   #3
Jeremy37
Member
 
Location: Montreal, Canada

Join Date: Feb 2011
Posts: 17
Default

I haven't dealt with RNA Seq data before... but isn't the difference just the non-coding portions of the mRNAs? I.e. 5' and 3' UTRs.

Mapping to coding sequences only would not include UTRs, whereas cDNA would.
Jeremy37 is offline   Reply With Quote
Old 07-05-2011, 02:49 AM   #4
sisch
Member
 
Location: Dusseldorf, Germany

Join Date: Jun 2011
Posts: 29
Default

It's just as Jeremy37 says. cDNA contains in addition to the coding parts also the UTRs.

Maybe there's just a huge bias towards UTRs when doing 454 sequencing.
For now i will be checking this phenomenon on different datasets.
sisch is offline   Reply With Quote
Old 07-05-2011, 05:02 AM   #5
Eric Fournier
Member
 
Location: Quebec, Canada

Join Date: Jul 2011
Posts: 21
Default

Quote:
Originally Posted by sisch View Post
It's just as Jeremy37 says. cDNA contains in addition to the coding parts also the UTRs.

Maybe there's just a huge bias towards UTRs when doing 454 sequencing.
For now i will be checking this phenomenon on different datasets.
A bias toward UTRs might arise from the protocol used to prepare your samples. For example, if you amplified your material using oligo-dT primers, you'll have a bias toward the 3'UTRs.
Eric Fournier is offline   Reply With Quote
Old 07-05-2011, 05:31 AM   #6
Jeremy37
Member
 
Location: Montreal, Canada

Join Date: Feb 2011
Posts: 17
Default

Sisch, I still don't understand the problem. Why do you think it's wrong that a 0.4 fraction of reads maps to CDS and 0.6 maps to cDNA? You don't think the UTRs could be that long?

I don't know what the average is, but some UTRs are very long. You could calculate the expected fraction by comparing the same genes in CCDS vs. Refseq. To match your numbers you would expect the UTRs to comprise about half as much sequence as the CDS (since 0.6 is 50% more than 0.4).
Jeremy37 is offline   Reply With Quote
Old 07-05-2011, 11:20 PM   #7
sisch
Member
 
Location: Dusseldorf, Germany

Join Date: Jun 2011
Posts: 29
Default

@Eric: I just checked the library protocol again. It was a random primed amplification.
@Jeremy37: It's pretty much, that I didn't really understand, why I had a so much better mapping to cDNA than to cds. Now on second thought, the mapping to UTRs could make up for the 20%. Maybe I got a bit distracted, that less than 40% mapped, so I looked for a flaw in either sequencing or processing.

Thank you all for your contributions. I consider this thread as solved with the following conclusion for future readers:

A better mapping to cDNA in contrast to cds has no further implications on the sequencing, as long as the better mapping can be explained by sequence information derived from the UTRs (which won't map on cds obviously)
sisch is offline   Reply With Quote
Reply

Tags
cdna, cds, contamination, mapping, rna seq

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 02:11 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2018, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO