Go Back   SEQanswers > Applications Forums > RNA Sequencing

Similar Threads
Thread Thread Starter Forum Replies Last Post
intersect (actually: filter) a gtf file with coordinates from a bed-file dietmar13 Bioinformatics 6 07-13-2017 05:20 AM
Problem generating counts with featureCount Joshua.Urrutia RNA Sequencing 10 11-24-2015 07:44 PM
Creating Transcriptome File for Use With BWA from GTF file and genomic fasta file PolPittacus7 Bioinformatics 4 07-17-2015 09:45 AM
what normalization should I do to compare featureCount outcome roll Bioinformatics 1 02-03-2014 04:40 AM
Count table from BAM file with custom gtf tamari Bioinformatics 3 09-05-2013 02:42 PM

Thread Tools
Old 03-02-2016, 06:56 AM   #1
Junior Member
Location: New York City

Join Date: Mar 2016
Posts: 3
Default featureCount with custom GTF file

Hi all:

My name is Haritz Irizar and I am a postdoctoral fellow at the Icahn School of Medicine at Mount Sinai.

Working with RNAseq data, our group routinely uses featurecounts to obtain both gene-level and exon-level count matrices from .bam files.

In our last analysis, and with the objective to obtain the read counts not only for the genes of the host but also for the bacteria of the microbiome from the RNAseq data obtained from mouse colon biopsies, we have built a “superGenome” reference composed of around 1500 microbial genomes and created a custom GTF annotations file for that superGenome.

After some problems, we have managed to do the alignment with STAR using both the mouse genome and the superGenome, but when we have tried to use featurecounts on the resulting .bam files, we get an error concerning the GTF file.

To build the count matrix, we have concatenated the mouse annotation GTF and the superGenome annotation GTF files and fed the concatenated GTF to featurecounts and this is the error we get for several lines of the concatenated GTF file that correspond to the microbial annotations:

Warning: the feature on the 1570797-th line has zero coordinate or zero lengths

Warning: the feature on the 1570798-th line has zero coordinate or zero lengths

Any idea of what could be wrong?

Could please any of you have a look at the error logs of the featurecounts run (attached in pdf format) and at the extract of the GTF annotation file for the microbial genomes (in .xml format)?

Thanks a lot,

Attached Files
File Type: xml SuperGenome_gtf_extract.xml (33.0 KB, 1 views)
File Type: pdf feature_counts_error_log.pdf (57.0 KB, 1 views)
haritz_irizar is offline   Reply With Quote
Old 03-02-2016, 07:17 AM   #2
Senior Member
Location: East Coast USA

Join Date: Feb 2008
Posts: 7,053

Can you post the output of

$ sed -n '1570797,1570798p' your_gtf_file
so we can see that is in those two lines that generates an error?
GenoMax is offline   Reply With Quote
Old 03-02-2016, 09:14 AM   #3
Junior Member
Location: New York City

Join Date: Mar 2016
Posts: 3

Hi GenoMax:

These are 2 of the lines (lines 1570797 and 1570798) of the concatenated GTF file that give an error:

SuperGenome custom exon 2422121283 2423917566 . + . gene_id "Leuconostoc"; transcript_id "Leuconostoc_trans_784"; exon_id "Leuconostoc citreum KM20, complete genome";
SuperGenome custom exon 2423927567 2425821065 . + . gene_id "Leuconostoc"; transcript_id "Leuconostoc_trans_785"; exon_id "Leuconostoc gelidum JB7, complete genome";
Thanks for your help,

haritz_irizar is offline   Reply With Quote

feature counts, gtf file, rnaseq

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 01:21 AM.

Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2020, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO