Problems with cuffmerge

Nnogueira · 10-27-2011, 09:35 AM

0

Hi everyone,

I'm trying to use cuffmerge but I get the following error message:

[Thu Oct 27 13:47:16 2011] Converting GTF files to SAM [FAILED] Error: gtf_to_sam not found on this system. Did you forget to include it in your PATH?

My command line is:

/storage/app/cufflinks-1.0.3/cufflinks-1.0.3.Linux_x86_64/cuffmerge -g /storage/data/natasha/gtf/miRNA_related_tracks_known_miRNAs_hg18.gtf -s /storage/data/natasha/chrs_fa/hg18_no_random_no_hap.fa assembles_cancer.txt

I've tested the gtf_to_sam program and it works fine.

Do I have do put something else in my command line? I've read the manual, but I couldn't find anything...

thanks in advance!

westerman · 10-27-2011, 11:03 AM

How did you test the gtf_to_sam program? Please give the command line you used to do the test.

ercfrtz · 11-01-2011, 10:57 AM

I have fixed the issue.

Nnogueira · 11-01-2011, 11:40 AM

Hi,

thanks for the replys! I've already fixed it. It was a problem with my bash file, nothing to do with cufflinks... sorry!

To answer the other reply, my command line for testing the gtf_to_sam program was

cufflinks-1.0.3.Linux_x86_64/gtf_to_sam transcripts.gtf SRR04006_gtf2sam.sam

and it worked fine.

thanks again for the replys!

westerman · 11-01-2011, 11:44 AM

Glad you got it fixed. Just to help out people who may read this topic. I suspect that gtf_to_sam was not in your PATH. Thus the fix to your bash file as well as using a given path (cufflinks-1.0.3.Linux_x86_64) to start up the program.

cewells · 01-02-2012, 09:58 AM

I am having the same issue, i.e.

"Error:gtf_to_sam not found on this system. Did you forget to include it in your PATH?"

I have included the path to gtf_to_sam in my .bashrc file PATH variable, but still I get this error.

Any thoughts? I appreciate your help --

Christina

jbrwn · 01-02-2012, 10:54 AM

Quote:

Originally Posted by cewells

I am having the same issue, i.e.

"Error:gtf_to_sam not found on this system. Did you forget to include it in your PATH?"

I have included the path to gtf_to_sam in my .bashrc file PATH variable, but still I get this error.

Any thoughts? I appreciate your help --

Christina

what's the location of gtf_to_sam? what's your $PATH? did you source your bashrc or log out and back in?

cewells · 01-02-2012, 11:12 AM

With help from Cole Trapnell, I pinpointed the problem.

In case anyone runs into the same issue, gtf_to_sam should be specified in your PATH as:

PATH=$PATH:/home/cufflinks/ <or wherever you've stored gtf_to_sam>

Then I used

. ~/.bashrc

to reload the .bashrc file without logging out.

-Christina

wangli · 05-30-2012, 08:10 AM

Dear cewells

I defined the path in the command line.
But when I utilized .~/.bashrc, it showed that it cannot find the file or directory.
I used ls -a to see the hidden file, but no .bashrc.
Do you have an idea of this problem?
Thanks!

westerman · 05-30-2012, 08:31 AM

Quote:

Originally Posted by wangli

Dear cewells

I defined the path in the command line.
But when I utilized .~/.bashrc, it showed that it cannot find the file or directory.
I used ls -a to see the hidden file, but no .bashrc.
Do you have an idea of this problem?
Thanks!

Did you edit your .bashrc or just put the path on the command line?

The basic idea is to
1) Edit your .bashrc or .bash_profile to have the new PATH in it.

2) Either logout/login or source a 'source' or a '.' of the .bashrc or .bash_profile in order to read the file in and make the PATH change.

I should point out that doing the above is basic UNIX 101. You will not get far unless you understand how your shell gets set up in the first place.

wanfahmi · 12-03-2012, 07:27 AM

Hi,

I got an error when I am at the stage of merge using cuffmerge. Below is my error

$ cuffmerge -g genes.gtf -s genome.fa -p 4 assemblies.txt

[Mon Dec 3 15:23:23 2012] Beginning transcriptome assembly merge
-------------------------------------------

[Mon Dec 3 15:23:23 2012] Preparing output location ./merged_asm/
Traceback (most recent call last):
File "/usr/local/cufflinks/cuffmerge", line 576, in <module>
sys.exit(main())
File "/usr/local/cufflinks/cuffmerge", line 538, in main
gtf_input_files = test_input_files(transfrag_list_file)
File "/usr/local/cufflinks/cuffmerge", line 268, in test_input_files
g = open(line,"r")
IOError: [Errno 2] No such file or directory: '1\tCufflinks\ttranscript\t89917\t91031\t1000\t-\t.\tgene_id "CUFF.1"; transcript_id "CUFF.1.1"; FPKM "0.6709083067"; frac "1.000000"; conf_lo "0.316487"; conf_hi "1.025329"; cov "2.502314";'

Please somebody help. Thank you!

westerman · 12-03-2012, 07:32 AM

Your 'assemblies.txt' file does not contain a list of files but rather contains data. At least that is my first guess.

wanfahmi · 12-03-2012, 07:50 AM

Hi Westerman,

Thanks for replying. do you mean the 'assemblies.txt' should have a list? Actually you are correct I guess as I did cat the all transcript.gtf. How should I do? Thanks

westerman · 12-03-2012, 08:01 AM

I do not do this too often and I am too busy at the moment to double-check but what you should have is a list of the individual files. E.g.,

cat >assemblies.txt
file1.gtf
file2.gtf
etc.

Not the contents of the files. The file names.

wanfahmi · 12-03-2012, 08:05 AM

Problems solved! thank you.

I did create a file that only contains the names not the contains of file. Its running.

Gonza · 09-13-2014, 12:09 PM

Hi all, I have a cuffmerge problem. It says "EOF marker is absent. The input is probably truncated". I have run tophat and cufflinks successfully. What does this mean??
Thanks.
G

[gonzalo@localhost trimmed_reads]$ cuffmerge -g genes.gtf -s genome.fa -p 20 assemblies.txt

[Sat Sep 13 12:33:25 2014] Beginning transcriptome assembly merge
-------------------------------------------

[Sat Sep 13 12:33:25 2014] Preparing output location ./merged_asm/
[Sat Sep 13 12:33:35 2014] Converting GTF files to SAM
[12:33:35] Loading reference annotation.
[12:33:37] Loading reference annotation.
[12:33:40] Loading reference annotation.
[12:33:43] Loading reference annotation.
[12:33:45] Loading reference annotation.
[12:33:48] Loading reference annotation.
[Sat Sep 13 12:33:51 2014] Quantitating transcripts
You are using Cufflinks v2.2.1, which is the most recent release.
Command line:
cufflinks -o ./merged_asm/ -F 0.05 -g genes.gtf -q --overhang-tolerance 200 --library-type=transfrags -A 0.0 --min-frags-per-transfrag 0 --no-5-extend -p 20 ./merged_asm/tmp/mergeSam_fileJRPl29
[bam_header_read] EOF marker is absent. The input is probably truncated.
[bam_header_read] invalid BAM binary header (this is not a BAM file).
File ./merged_asm/tmp/mergeSam_fileJRPl29 doesn't appear to be a valid BAM file, trying SAM...
[12:33:51] Loading reference annotation.
[12:33:57] Inspecting reads and determining fragment length distribution.
Processed 28060 loci.
Map Properties:
Normalized Map Mass: 154834.00
Raw Map Mass: 154834.00
Fragment Length Distribution: Truncated Gaussian (default)
Default Mean: 200
Default Std Dev: 80
[12:33:59] Assembling transcripts and estimating abundances.
Processed 28060 loci.
[Sat Sep 13 12:34:11 2014] Comparing against reference file genes.gtf
You are using Cufflinks v2.2.1, which is the most recent release.
[Sat Sep 13 12:34:34 2014] Comparing against reference file genes.gtf
You are using Cufflinks v2.2.1, which is the most recent release.

Brian Bushnell · 09-13-2014, 12:38 PM

That message is a bug in Samtools; don't worry about it.

Gonza · 09-13-2014, 06:40 PM

Thanks. I ignored it and successfully run tuxedo. However, when loading cuffdiff to R i keep getting this error. I have no idea what this is. Any thiughts on solutions? thanks...

> cuff_data <- readCufflinks()
Creating database /Users/gonzalovillarino/Desktop/05_cuffdiff/cuffData.db
Reading Run Info File /Users/gonzalovillarino/Desktop/05_cuffdiff/run.info
Writing runInfo Table
Reading Read Group Info /Users/gonzalovillarino/Desktop/05_cuffdiff/read_groups.info
Writing replicates Table
Reading /Users/gonzalovillarino/Desktop/05_cuffdiff/genes.fpkm_tracking
Error in scan(file, what, nmax, sep, dec, quote, skip, nlines, na.strings, :
line 1305 did not have 17 elements

> cuff_data
Error: object 'cuff_data' not found
CuffSet instance with:
0 samples
0 genes
0 isoforms
0 TSS
0 CDS
0 promoters
0 splicing
0 relCDS

Nanu · 10-14-2014, 04:43 AM

When I am running the cuffmerge then i got the following error. I mentioned the .gtf file name in it which was created by cufflinks

[Tue Oct 14 17:08:29 2014] Beginning transcriptome assembly merge
-------------------------------------------

[Tue Oct 14 17:08:29 2014] Preparing output location ./merged_asm/
Warning: no reference GTF provided!
[Tue Oct 14 17:08:31 2014] Converting GTF files to SAM
[FAILED]
Error: gtf_to_sam not found on this system. Did you forget to include it in your PATH?

Rashedul Islam · 12-09-2014, 02:50 PM

I have solved the problem by adding this line to the .hashrc file.

"export PATH=$PATH:/home/cufflinks-2.2.1.Linux_x86_64"

Similar Threads
Thread	Thread Starter	Forum	Replies	Last Post
Cuffmerge Error	ercfrtz	Bioinformatics	12	05-19-2014 10:02 PM
cuffmerge failed	wall_y	Bioinformatics	4	07-12-2012 07:25 AM
cuffmerge problems	ccstaats	Bioinformatics	0	02-02-2012 05:28 AM
Questions about cuffmerge	Jolin	Bioinformatics	0	10-04-2011 01:46 AM
cuffmerge fail	papori	Bioinformatics	0	07-31-2011 04:01 PM

10-27-2011, 09:35 AM	#1
Nnogueira Junior Member Location: Brazil Join Date: Oct 2011 Posts: 9	Problems with cuffmerge 0 Hi everyone, I'm trying to use cuffmerge but I get the following error message: [Thu Oct 27 13:47:16 2011] Converting GTF files to SAM [FAILED] Error: gtf_to_sam not found on this system. Did you forget to include it in your PATH? My command line is: /storage/app/cufflinks-1.0.3/cufflinks-1.0.3.Linux_x86_64/cuffmerge -g /storage/data/natasha/gtf/miRNA_related_tracks_known_miRNAs_hg18.gtf -s /storage/data/natasha/chrs_fa/hg18_no_random_no_hap.fa assembles_cancer.txt I've tested the gtf_to_sam program and it works fine. Do I have do put something else in my command line? I've read the manual, but I couldn't find anything... thanks in advance!

11-01-2011, 10:57 AM	#3
ercfrtz Member Location: Iowa Join Date: Aug 2010 Posts: 23	I have fixed the issue. Last edited by ercfrtz; 11-01-2011 at 11:00 AM.

01-02-2012, 09:58 AM	#6
cewells Junior Member Location: South Carolina Join Date: Jul 2011 Posts: 3	clarification I am having the same issue, i.e. "Error:gtf_to_sam not found on this system. Did you forget to include it in your PATH?" I have included the path to gtf_to_sam in my .bashrc file PATH variable, but still I get this error. Any thoughts? I appreciate your help -- Christina

10-27-2011, 11:03 AM	#2
westerman Rick Westerman Location: Purdue University, Indiana, USA Join Date: Jun 2008 Posts: 1,104	How did you test the gtf_to_sam program? Please give the command line you used to do the test.

11-01-2011, 11:40 AM	#4
Nnogueira Junior Member Location: Brazil Join Date: Oct 2011 Posts: 9	Hi, thanks for the replys! I've already fixed it. It was a problem with my bash file, nothing to do with cufflinks... sorry! To answer the other reply, my command line for testing the gtf_to_sam program was cufflinks-1.0.3.Linux_x86_64/gtf_to_sam transcripts.gtf SRR04006_gtf2sam.sam and it worked fine. thanks again for the replys!

11-01-2011, 11:44 AM	#5
westerman Rick Westerman Location: Purdue University, Indiana, USA Join Date: Jun 2008 Posts: 1,104	Glad you got it fixed. Just to help out people who may read this topic. I suspect that gtf_to_sam was not in your PATH. Thus the fix to your bash file as well as using a given path (cufflinks-1.0.3.Linux_x86_64) to start up the program.

01-02-2012, 11:12 AM	#8
cewells Junior Member Location: South Carolina Join Date: Jul 2011 Posts: 3	With help from Cole Trapnell, I pinpointed the problem. In case anyone runs into the same issue, gtf_to_sam should be specified in your PATH as: PATH=$PATH:/home/cufflinks/ <or wherever you've stored gtf_to_sam> Then I used . ~/.bashrc to reload the .bashrc file without logging out. -Christina

05-30-2012, 08:10 AM	#9
wangli Member Location: Texas Join Date: Apr 2012 Posts: 48	Dear cewells I defined the path in the command line. But when I utilized .~/.bashrc, it showed that it cannot find the file or directory. I used ls -a to see the hidden file, but no .bashrc. Do you have an idea of this problem? Thanks!

12-03-2012, 07:27 AM	#11
wanfahmi Member Location: North Sea Join Date: Apr 2008 Posts: 34	Hi, I got an error when I am at the stage of merge using cuffmerge. Below is my error $ cuffmerge -g genes.gtf -s genome.fa -p 4 assemblies.txt [Mon Dec 3 15:23:23 2012] Beginning transcriptome assembly merge ------------------------------------------- [Mon Dec 3 15:23:23 2012] Preparing output location ./merged_asm/ Traceback (most recent call last): File "/usr/local/cufflinks/cuffmerge", line 576, in <module> sys.exit(main()) File "/usr/local/cufflinks/cuffmerge", line 538, in main gtf_input_files = test_input_files(transfrag_list_file) File "/usr/local/cufflinks/cuffmerge", line 268, in test_input_files g = open(line,"r") IOError: [Errno 2] No such file or directory: '1\tCufflinks\ttranscript\t89917\t91031\t1000\t-\t.\tgene_id "CUFF.1"; transcript_id "CUFF.1.1"; FPKM "0.6709083067"; frac "1.000000"; conf_lo "0.316487"; conf_hi "1.025329"; cov "2.502314";' Please somebody help. Thank you!

12-03-2012, 07:32 AM	#12
westerman Rick Westerman Location: Purdue University, Indiana, USA Join Date: Jun 2008 Posts: 1,104	Your 'assemblies.txt' file does not contain a list of files but rather contains data. At least that is my first guess.

12-03-2012, 07:50 AM	#13
wanfahmi Member Location: North Sea Join Date: Apr 2008 Posts: 34	Hi Westerman, Thanks for replying. do you mean the 'assemblies.txt' should have a list? Actually you are correct I guess as I did cat the all transcript.gtf. How should I do? Thanks

12-03-2012, 08:01 AM	#14
westerman Rick Westerman Location: Purdue University, Indiana, USA Join Date: Jun 2008 Posts: 1,104	I do not do this too often and I am too busy at the moment to double-check but what you should have is a list of the individual files. E.g., cat >assemblies.txt file1.gtf file2.gtf etc. Not the contents of the files. The file names.

12-03-2012, 08:05 AM	#15
wanfahmi Member Location: North Sea Join Date: Apr 2008 Posts: 34	Problems solved! thank you. I did create a file that only contains the names not the contains of file. Its running.

09-13-2014, 12:09 PM	#16
Gonza Member Location: Ithaca, NY Join Date: Mar 2013 Posts: 78	Hi all, I have a cuffmerge problem. It says "EOF marker is absent. The input is probably truncated". I have run tophat and cufflinks successfully. What does this mean?? Thanks. G [gonzalo@localhost trimmed_reads]$ cuffmerge -g genes.gtf -s genome.fa -p 20 assemblies.txt [Sat Sep 13 12:33:25 2014] Beginning transcriptome assembly merge ------------------------------------------- [Sat Sep 13 12:33:25 2014] Preparing output location ./merged_asm/ [Sat Sep 13 12:33:35 2014] Converting GTF files to SAM [12:33:35] Loading reference annotation. [12:33:37] Loading reference annotation. [12:33:40] Loading reference annotation. [12:33:43] Loading reference annotation. [12:33:45] Loading reference annotation. [12:33:48] Loading reference annotation. [Sat Sep 13 12:33:51 2014] Quantitating transcripts You are using Cufflinks v2.2.1, which is the most recent release. Command line: cufflinks -o ./merged_asm/ -F 0.05 -g genes.gtf -q --overhang-tolerance 200 --library-type=transfrags -A 0.0 --min-frags-per-transfrag 0 --no-5-extend -p 20 ./merged_asm/tmp/mergeSam_fileJRPl29 [bam_header_read] EOF marker is absent. The input is probably truncated. [bam_header_read] invalid BAM binary header (this is not a BAM file). File ./merged_asm/tmp/mergeSam_fileJRPl29 doesn't appear to be a valid BAM file, trying SAM... [12:33:51] Loading reference annotation. [12:33:57] Inspecting reads and determining fragment length distribution. Processed 28060 loci. Map Properties: Normalized Map Mass: 154834.00 Raw Map Mass: 154834.00 Fragment Length Distribution: Truncated Gaussian (default) Default Mean: 200 Default Std Dev: 80 [12:33:59] Assembling transcripts and estimating abundances. Processed 28060 loci. [Sat Sep 13 12:34:11 2014] Comparing against reference file genes.gtf You are using Cufflinks v2.2.1, which is the most recent release. [Sat Sep 13 12:34:34 2014] Comparing against reference file genes.gtf You are using Cufflinks v2.2.1, which is the most recent release.

09-13-2014, 12:38 PM	#17
Brian Bushnell Super Moderator Location: Walnut Creek, CA Join Date: Jan 2014 Posts: 2,707	That message is a bug in Samtools; don't worry about it.

09-13-2014, 06:40 PM	#18
Gonza Member Location: Ithaca, NY Join Date: Mar 2013 Posts: 78	Thanks. I ignored it and successfully run tuxedo. However, when loading cuffdiff to R i keep getting this error. I have no idea what this is. Any thiughts on solutions? thanks... > cuff_data <- readCufflinks() Creating database /Users/gonzalovillarino/Desktop/05_cuffdiff/cuffData.db Reading Run Info File /Users/gonzalovillarino/Desktop/05_cuffdiff/run.info Writing runInfo Table Reading Read Group Info /Users/gonzalovillarino/Desktop/05_cuffdiff/read_groups.info Writing replicates Table Reading /Users/gonzalovillarino/Desktop/05_cuffdiff/genes.fpkm_tracking Error in scan(file, what, nmax, sep, dec, quote, skip, nlines, na.strings, : line 1305 did not have 17 elements > cuff_data Error: object 'cuff_data' not found CuffSet instance with: 0 samples 0 genes 0 isoforms 0 TSS 0 CDS 0 promoters 0 splicing 0 relCDS

10-14-2014, 04:43 AM	#19
Nanu Member Location: New Delhi Join Date: Sep 2014 Posts: 30	When I am running the cuffmerge then i got the following error. I mentioned the .gtf file name in it which was created by cufflinks [Tue Oct 14 17:08:29 2014] Beginning transcriptome assembly merge ------------------------------------------- [Tue Oct 14 17:08:29 2014] Preparing output location ./merged_asm/ Warning: no reference GTF provided! [Tue Oct 14 17:08:31 2014] Converting GTF files to SAM [FAILED] Error: gtf_to_sam not found on this system. Did you forget to include it in your PATH?

12-09-2014, 02:50 PM	#20
Rashedul Islam Junior Member Location: Canada Join Date: May 2012 Posts: 1	I have solved the problem by adding this line to the .hashrc file. "export PATH=$PATH:/home/cufflinks-2.2.1.Linux_x86_64"