RNAseq error while running sem-calculate-expression

Question

[shuksi1984]

Hi,

I ran following command:

/path/to/rsem/rsem-calculate-expression -p 2 --output-genome-bam -star --star-path /path/to

/STAR_index_files --paired-end path/to/SRR925687_1.fastq path/to/SRR925687_2.fastq human.refseq.gff.gtf path/to/RNA.test

I got following error msg:

dFilesIn SRR925687_1.fastq SRR925687_2.fastq" failed! Plase check if you provide correct parameters/options for the pipeline!

STAR index folder consists of following files:

rwxrwxr-x 2 stuser stuser 4096 Apr 17 18:16 ./

drwxrwxr-x 7 stuser stuser 4096 Apr 18 11:12 ../

-rw-r--r-- 1 root root 1194 Apr 9 18:37 chrLength.txt

-rw-r--r-- 1 root root 3532 Apr 9 18:37 chrNameLength.txt

-rw-r--r-- 1 root root 2338 Apr 9 18:37 chrName.txt

-rw-r--r-- 1 root root 2108 Apr 9 18:37 chrStart.txt

-rw-r--r-- 1 root root 51816787 Apr 9 19:41 exonGeTrInfo.tab

-rw-r--r-- 1 root root 23213627 Apr 9 19:41 exonInfo.tab

-rw-r--r-- 1 root root 380243 Apr 9 19:41 geneInfo.tab

-rw-r--r-- 1 root root 3202741093 Apr 9 21:34 Genome

-rw-r--r-- 1 root root 858 Apr 9 21:25 genomeParameters.txt

-rw-r--r-- 1 root root 12693443564 Apr 9 22:47 SA

-rw-r--r-- 1 root root 1565873619 Apr 9 22:54 SAindex

-rw-r--r-- 1 root root 9414412 Apr 9 19:41 sjdbInfo.txt

-rw-r--r-- 1 root root 10724637 Apr 9 19:41 sjdbList.fromGTF.out.tab

-rw-r--r-- 1 root root 10723973 Apr 9 19:41 sjdbList.out.tab

-rw-r--r-- 1 root root 7215534 Apr 9 19:41 transcriptInfo.tab

I have provided correct path to the files generated by STAR indexing. Dont understand the problem.

[Devon Ryan]

It's likely that human.refseq.gff.gtf is an incorrect reference name. Did you use that with rsem-prepare-reference? BTW, your error message looks truncated.

[shuksi1984]

Yes, human.refseq.gff.gtf was generated with rsem-prepare-reference. The same file was used during indexing with STAR.

[shuksi1984]

Following shows the complete error.

sh: 1: /data/ngs/mona/RNAseq/genome/genome_file/STAR: not found
"/data/ngs/mona/RNAseq/genome/genome_file/STAR --genomeDir .  --outSAMunmapped Within  --outFilterType BySJout  --outSAMattributes NH HI AS NM MD  --outFilterMultimapNmax 20  --outFilterMismatchNmax 999  --outFilterMismatchNoverLmax 0.04  --alignIntronMin 20  --alignIntronMax 1000000  --alignMatesGapMax 1000000  --alignSJoverhangMin 8  --alignSJDBoverhangMin 1  --sjdbScore 1  --runThreadN 2  --genomeLoad NoSharedMemory  --outSAMtype BAM Unsorted  --quantMode TranscriptomeSAM  --outSAMheaderHD \@HD VN:1.4 SO:unsorted  --outFileNamePrefix RNA.test.temp/RNA.test  --readFilesIn SRR925687_1.fastq SRR925687_2.fastq" failed! Plase check if you provide correct parameters/options for the pipeline!
stuser@beta:/data/ngs/mona/RNAseq/genome$ cd ../../../