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I am trying to check the quality of RNA-Seq data from Illumina using fastqc in snakemake in a conda environment. I get the error "Multiple run or shell keywords in rule run_fastqc".

Snakefile:

rule run_fastqc:
    input:
        "/[RNA fastq file path]/well01_S001_L001_R1_001.fastq.gz"
    output:
        "./[out path]/well01_S001_L001_R1_001.fastqc.html",
        "./[out path]/well01_S001_L001_R1_001.fastqc.zip"
    wrapper:
        "0.79.0/bio/fastqc"
    shell:
        "fastqc {input} --outdir=./[out path]/"

I've also tried:

...
    shell:
        "fastqc {input} -o ./[out path]/"
...
    shell:
        "fastqc -o ./[out path]/ {input}"
...
    shell:
        "fastqc {input}"
        "fastqc -o ./[out path]/"
...
    shell:
        "fastqc {input} > -o ./[out path]/"

Code:

snakemake --cores 1 well01_S001_L001_R1_001.fastqc.html well01_S001_L001_R1_001.fastqc.zip

Output:

SyntaxError in line 9 of /[path to Snakefile]/Snakefile:
Multiple run or shell keywords in rule run_fastqc. (Snakefile, line 9)

When I run fastqc alone (not in snakemake), it works and I get the two output files. How can I get snakemake to work?

Thank you for your time and help!

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From a quick look at the manual, it looks like you are supposed to use just the wrapper, not combine the wrapper with a shell. Can you show me an example of such combined usage?

The error seems to suggest that the wrapper already has a shell directive that you shell conflicts with.

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  • $\begingroup$ Thank you! In Output, I also had to remove the "./[out path]/" before the output file name, I guess the "--outdir" in Shell is sufficient. I got the files but still received the error: "Job Missing files after 5 seconds: well01_S001_L001_R1_001.fastqc.html well01_S001_L001_R1_001.fastqc.zip This might be due to filesystem latency. If that is the case, consider to increase the wait time with --latency-wait. Job id: 0 completed successfully, but some output files are missing. 0" and I also waited for 60s and got the same error. $\endgroup$
    – Swietenia
    Nov 11 at 21:06

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