Input files not found

If only no file, only one input file , or only read one and not read two is picked up then something is wrong with your input file declaration

  1. The path must be enclosed in quotes (' or ")
  2. The path must have at least one * wildcard character. This is even if you are only running one paired end sample.
  3. When using the pipeline with paired end data, the path must use {1,2} or {R1,R2} notation to specify read pairs.
  4. If you are running Single end data make sure to specify --singleEnd

If the pipeline can't find your files then you will get the following error

ERROR ~ Cannot find any reads matching: *{1,2}.fastq.gz

Note that if your sample name is "messy" then you have to be very particular with your glob specification. A file name like L1-1-D-2h_S1_L002_R1_001.fastq.gz can be difficult enough for a human to read. Specifying *{1,2}*.gz wont work give you what you want Whilst *{R1,R2}*.gz will.

Data organization

The pipeline can't take a list of multiple input files - it takes a glob expression. If your input files are scattered in different paths then we recommend that you generate a directory with symlinked files. If running in paired end mode please make sure that your files are sensibly named so that they can be properly paired. See the previous point.

Image cannot be built

Sometimes, mksquashfs cannot be found on the login node or workstation that you intend to use, thus the Singularity Image build fails unfortunately. See below code snippet that shows such a typical failure:

Caused by:
  Failed to pull singularity image
  command: singularity pull --name nfcore-rnaseq-1.3.img docker://nfcore/rnaseq:1.3 > /dev/null
  status : 255
  .....
    INFO:    Creating SIF file...
    FATAL:   Unable to pull docker://nfcore/rnaseq:1.3: While searching for mksquashfs: exec: "mksquashfs": executable file not found in $PATH

If this is the case, please install mksquashfs or ask your IT department to install the package for you.

Cannot find input files when using Singularity

Depending on how you install Singularity on your system several reoccurring issues have been reported. Typically these result in error messages such as these:


Command error:
  �[91mERROR  : Failed to resolve path to /home/bla/singularity/mnt/container: No such file or directory
  �[0m�[31mABORT  : Retval = 255
  �[0m

You can verify if this is indeed a problem by using a Singularity Shell to access your container, and to check whether the required paths are available inside your container:

singularity shell /path/to/work/singularity/imagename.simg
ls -la /path/to/work
ls -la /path/to/network_storage

If any of these ls -la commands returns a Not Found error, you will need to set/update Singularity Bind Paths on your system.

The Singularity installation requires certain host paths to be bound. Please see Singularity Bind Paths Documentation for a more detailed explanation. In many cases this can be resolved by adding these paths to your /etc/singularity/singularity.conf as highlighted in the documentation:

bind path = /beegfs/work/
bind path = /scratch
bind path = /gpfs
bind path = /home

Alternatively, you can also add Singularity Bind Paths to your Nextflow call, e.g. using autoMounts and/or runOptions in the Singularity scope

Warning about sticked on revision

If you get a warning like the following:

Project nf-core/eager currently is sticked on revision: dev -- you need to specify explicitly a revision with the option -r to use it

This is a nextflow error, with less commonly seen terminology. What this means is that you have multiple versions of nf-core/eager pulled (e.g. 2.0.0, 2.1.0, 2.1.1, dev etc.). When you have multiple versions, you must always specify which one you want to use - there is no 'default'. Therefore, with every nextflow run nf-core/<PIPELINE> command you must always indicate which version with -r.

For example:

nextflow run nf-core/eager -r 2.1.0 --reads '/<path>/<to>/data/*_{R1,R2}_*.fq.gz' <...>

Specifying the version of the run you are using is highly recommended, as it helps in full reproducibility. In the sense that if you explicitly record the whole command with the version for your publication or internal reports, then anyone who wants to check your work can use the exact version you used (including all internal tools).

You can see more information on the nextflow documentation here.

Extra resources and getting help

If you still have an issue with running the pipeline then feel free to contact us via the Slack channel or by opening an issue in the respective pipeline repository on GitHub asking for help.

If you have problems that are directly related to Nextflow and not our pipelines or the nf-core framework tools then check out the Nextflow gitter channel or the google group.