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Old 11-27-2013, 10:09 AM   #1
komalsrathi
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Location: Philadelphia, PA

Join Date: Aug 2013
Posts: 14
Default Cuffmerge output with missing strand information, htseq-count error

Hi everyone,

I used cuffmerge to obtain a merged.gtf file but there are 3700 entries in my merged.gtf file that contain '.' in the strand field instead of '+' or '-'. I checked the entries that do not have a strand information and each of these have a class-code "u" which stands for unknown.
I want to use this merged.gtf file in my htseq-count command to get the counts (I eventually plan to do a differential gene expression analysis). When I use htseq-count using merged.gtf I get the following error:

Quote:
Error occured when processing GFF file (line 360833 of file ./merged_asm/merged.gtf):
Feature XLOC_003190 at chr1:[1285003,1285358)/. does not have strand information but you are running htseq-count in stranded mode. Use '--stranded=no'.
[Exception type: SystemExit, raised in count.py:59]
I know the error is due to missing strand information. I have two options here:

1) Use --stranded=no
2) Remove all the entries corresponding to missing strand information

What do you guys suggest I do? Do you think removing entries with missing information is better than using --stranded=no? I am worried that using --stranded=no will affect the resulting counts, right? Are there any other suggestions?

FYI, I have posted this question on Biostars too:http://www.biostars.org/p/87578/
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Komal Rathi
Bioinformatics Application Developer
University of Pennsylvania

Last edited by komalsrathi; 11-27-2013 at 12:21 PM.
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Old 11-27-2013, 02:00 PM   #2
dpryan
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Location: Freiburg, Germany

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Posts: 3,476
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Were the libraries directional to begin with and did you tell cufflinks that?
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