SEQanswers

Go Back   SEQanswers > Bioinformatics > Bioinformatics



Similar Threads
Thread Thread Starter Forum Replies Last Post
Cufflinks' estimated fragment length mean and standard deviation d f Bioinformatics 12 01-18-2014 10:32 AM
CuffLinks/CuffDiff frag length for single-end reads mbblack Bioinformatics 2 12-17-2011 01:47 PM
cufflinks detecting single-end when paired-end used dnusol Bioinformatics 8 02-02-2011 08:43 AM
Does Cufflinks support single-end and paired end data together ? ersenkavak Bioinformatics 1 10-22-2010 07:26 AM
definition of "fragment" in FPKM in single end reads thinkRNA Bioinformatics 1 06-25-2010 06:00 AM

Reply
 
Thread Tools
Old 09-13-2011, 10:53 PM   #1
marcho
Junior Member
 
Location: Sweden

Join Date: Sep 2011
Posts: 5
Default Cufflinks - fragment mean length for single-end reads

Hi,

so this is probably going to be a very naive question, but here goes anyway:

I have just started using cufflinks - the dataset is single-end reads from the HiSeq2000 platform with a length of 100bp (105 if you count the adapter, which has already been removed).

I noticed a dramatic difference in FPKM value estimates between the 'default settings' for fragment length man (-m) and standard length deviation (-s). Apparently, cufflinks is not able to figure out the correct values from the data. But what has to go in there, exactly? And what program would be best suited to give me those estimates? I used the Picard toolbox to get mean fragment length (100, surprise surprise) - but standard deviation is not determined there.

Also, while I would guess that 100 is the value to put in, I used -q 15 in the bwa assembly, which performed quite a bit if soft clipping on some reads. Does that factor in in any way?

Thanks for any helpful comments!

/Marc
marcho is offline   Reply With Quote
Old 09-28-2011, 12:13 PM   #2
slavailn
Junior Member
 
Location: Lethbridge

Join Date: Mar 2011
Posts: 8
Default

Hi, Mark!
I'm facing same kind of problem with single-end reads. Have you been able to resolve this issue?

Thanks!
Slava.
slavailn is offline   Reply With Quote
Old 09-28-2011, 12:28 PM   #3
Jon_Keats
Senior Member
 
Location: Phoenix, AZ

Join Date: Mar 2010
Posts: 279
Default

This is a bit of a guess but my best guess is to leave these blank (ie. don't put in the command) for single end reads. Check the cufflinks output on stdout, I'm betting if figures out these are single end reads, and runs as such were those two options are not used.
Jon_Keats is offline   Reply With Quote
Old 09-28-2011, 01:48 PM   #4
slavailn
Junior Member
 
Location: Lethbridge

Join Date: Mar 2011
Posts: 8
Default

Thanks for reply, Jon!

The program runs with no errors and produces expected output files, but I'm a bit worried about Read Type shown as 0 bp single end? While it should be 62 bp. Also I'm not sure if I can leave Mean and Std Dev at defaults, considering that I had 62bp + 7 barcode + adapter. Would this influence the results?

Upper Quartile: 252.00
> Number of Multi-Reads: 508671 (with 1031383 total hits)
> Read Type: 0bp single-end
> Fragment Length Distribution: Truncated Gaussian (default)
> Default Mean: 200
> Default Std Dev: 80
[14:36:12] Modeling fragment count overdispersion.
[14:36:13] Calculating initial abundance estimates for multi-read correction.
> Processed 31353 loci. [*************************] 100%
[14:55:46] Testing for differential expression and regulation in locus.
> Processed 31353 loci. [*************************] 100%
slavailn is offline   Reply With Quote
Reply

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 04:05 PM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2021, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO