After running cufflink for two samples of RNA-Seq data, I used cuffmerge and executed the following command
cuffmerge -p 8 -g <directory>/<gff file> -s <directory>/<refernec fasta file> <directory>/assemblies.txt
assemblies.txt contains the
But I am getting following error
[Sun Jul 13 14:44:46 2014] Beginning transcriptome assembly merge ------------------------------------------- [Sun Jul 13 14:44:46 2014] Preparing output location ./merged_asm/ [Sun Jul 13 14:46:04 2014] Converting GTF files to SAM [14:46:04] Loading reference annotation. [14:47:10] Loading reference annotation. [14:48:06] Loading reference annotation. [14:49:23] Loading reference annotation. [14:50:29] Loading reference annotation. [14:51:44] Loading reference annotation. [Sun Jul 13 14:52:46 2014] Quantitating transcripts Warning: Your version of Cufflinks is not up-to-date. It is recommended that you upgrade to Cufflinks v2.2.1 to benefit from the most recent features and bug fixes (http://cufflinks.cbcb.umd.edu). Command line: cufflinks -o ./merged_asm/ -F 0.05 -g <directory>/<gff file> -q --overhang-tolerance 200 --library-type=transfrags -A 0.0 --min-frags-per-transfrag 0 --no-5-extend -p 8 ./merged_asm/tmp/mergeSam_file3DKFYb [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_file3DKFYb doesn't appear to be a valid BAM file, trying SAM... [14:52:49] Loading reference annotation. Error parsing strand (?) from GFF line: IWGSC_CSS_1DS_scaff_731014 . repeat_region 1 174 . ? .Name=trf;class=trf;repeat_consensus=ATTGGTATAGAACGCATGAAGAAACTCCATACAGATGGATCTTTAGACTCACTCAATTATGAAAAAATTGAGACATGCAAACCATGTCT;type=Tandem repeats [FAILED] Error: could not execute cufflinks
Thanks a lot. Yes, I already started running the script using GTF file.
sbdk82 whenever you figure out the solution to your question before others, please post it here as an answer or accept an answer that other people post. So that people can focus on other 'open' questions. Thanks!
Yes, I was about to do that but the script was running and I was not sure if using GTF file solved that issue.
I am new in Rna-seq... I am trying to run the cuffmerge but ir is giving me some error... Please, could you help me...
It seems there is some error in the GTF file. Please check if it is the correct one.
Hi sbdk82 and komal.rathi;
I checked the gtf file too. I changed my gtf file also and tried once again but still same error its giving.
assembly.txtcontain all the transcript.gtf files? Did you run tophat and cufflinks before running cuffmerge? Try sorting all the SAM/BAM files before running cufflinks.
Yeah, I got the output from tophat and cufflinks, and the
assembly.txtfile has all the
transcript.gtffiles. If I run
cuffmergethe following error has occurred..
nikhilvgbt Sorry for the delayed response. Did you get things worked out?
Yeah, it worked for me. The problem in the bam file is just because there is no proper order of Chromosomes in Reference genome and Gtf file. Thank you komal.rathi
and komal.rathi, how did you solve this error? So that other people may have an answer for this.