rsem-calculate-expression error message

Question

[walker]

I'm trying to calculate exppression from raw fastq, and I am getting error when rsem finish STAR alignment. here is my sh script:

fastp -i 1.fastq.gz -I 2.fastq.gz -o clean_1.fq.gz -O clean_2.fq.gz -j RNA.fastp.json -h RNA.fastp.html
   RSEM-1.3.0/bin/rsem-calculate-expression -p 2 --paired-end --phred33-quals --star --star-path PATH/TO/STAR --star-gzipped-read-file --star-output-genome-bam --estimate-rspd --append-names clean_1.fq.gz  clean_2.fq.gz PATH/TO/reference PATH/TO/2.RSEMstar

and here is my error message:

 ..... started STAR run
 ..... loading genome
 ..... started mapping
..... finished successfully

     "rsem-parse-alignments RSEM/grch37r87_STAR PATH/TO/2.RSEMstar.temp/2.RSEMstar PATH/TO/2.RSEMstar.stat/2.RSEMstar PATH/TO/2.RSEMstar.temp/2.RSEMstar.bam 3 -tag XM" failed! Plase check if you provide correct parameters/options for the pipeline!

anyone knows why and can help solve this questions? Thanks

[Matteo Schiavinato]

Is it normal that you have the same file specified in two arguments? -o clean_2.fq.gz -O clean_2.fq.gz

[walker]

Thanks for your answer and sorry for my mistake I write on this question! Acturally, -o clean_1.fq.gz -O clean_2.fq.gz is what i write in my script And I correct it in my question,Thanks!

[Matteo Schiavinato]

You also declare twice the --star argument in RSEM. That shouldn't be a problem, but since it seems that you're not super-solid on your command writing, perhaps check if every file you declare is in the folder you want it to be, check if every tag you added is exactly as it should be, and stuff like that (it's the cause of 99% of the errors in the bioinformatics world, regardless of the experience!).

[mrpetzel96]

Did you find a solution? I am facing the same issue

Answer 1

[Devon Ryan]

clean_1.fq.gz isn't being created by fastp, since you specified read 2 twice.

[walker]

Thanks for your answer and sorry for my mistake I write on this question! Acturally, -o clean_1.fq.gz -O clean_2.fq.gz is what i write in my script