Question: featureCount not processing reads
0
gravatar for amoltej
5.0 years ago by
amoltej90
Australia
amoltej90 wrote:

Dear All, 

I am trying to use featurecounts for the first time. I am following exactly as mentioned in the Rsubread manual. 

featureCounts(files='Aligned.nsortedByCoord.out.bam', annot='cuffcmp.combined.gtf', isGTFAnnotationFile=TRUE,isPairedEnd=TRUE)

after that the outcome looks like this

 

Processing Bcyt/Aligned.sortedByCoord.out.bam  ...

Warning: the feature on the 4366-th line has zero coordinate or zero lengths


Warning: the feature on the 26925-th line has zero coordinate or zero lengths


Warning: the feature on the 36138-th line has zero coordinate or zero lengths


Warning: the feature on the 38896-th line has zero coordinate or zero lengths


Warning: the feature on the 49896-th line has zero coordinate or zero lengths


Warning: the feature on the 51284-th line has zero coordinate or zero lengths


Warning: the feature on the 54658-th line has zero coordinate or zero lengths


Warning: the feature on the 55995-th line has zero coordinate or zero lengths

There are 84237 features loaded from the annotation file.
The 84237 features are sorted.
Number of chromosomes included in the annotation is 4940
The 0-th thread processed 0 reads
Number of fragments mapped to the features is: 0
Time cost = 1.6 seconds

 

It seems nothing is happening because there are zero processed reads. 

I am not sure what has went wrong? also please tell me more about the featurecount out come? how to read the file??

Thank you in advance.

Amol

rna-seq rsubread featurecounts • 1.5k views
ADD COMMENTlink modified 5.0 years ago by geek_y10k • written 5.0 years ago by amoltej90
0
gravatar for geek_y
5.0 years ago by
geek_y10k
Barcelona
geek_y10k wrote:

from the file name (cuffcmp.combined.gtf), I could see that you are using cufflinks generated gtf file.

Will the subreads package works with cufflinks annotation file ? I would suggest first try to use the standard annotations ( if available for your genome) using getInBuiltAnnotation and see if it works.

But I would like to know why you are using the cufflinks gtf file to count the reads ? whats your goal ?

ADD COMMENTlink written 5.0 years ago by geek_y10k
Hey Geek_y thanks The organism that I am working on does not have standard annotated genome. I have made my own gtf file using scipio program and the closely related organism's protein sequences. When I visualized my BAM file, genome scaffold file and gtf file in seqmonk program I found out that there are so many reads which does not have annotation in my scipio gtf file. That's why I generated combined gtf file using cuffcompare which has all the annotations generated by cufflinks. I have just tried scipio gtf file and error is still same with more warnings.
ADD REPLYlink written 5.0 years ago by amoltej90
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