nanopolish | Signal-level algorithms for MinION data | Genomics library

The rule that fails is rule guppyplex, which looks for an input in the form of {FASTQ_PATH}/{{barcode}}.

Looks like the wildcard {barcode} is filled with barcode49/barcode49.consensus.fasta, which happened because of two reasons I think:

First (and most important): The workflow does not find a better way to produce the final output. In rule catFasta, you give an input file which is never described as an output in your workflow. The rule minion has the directory as an output, but not the file, and it is not perfectly clear for the workflow where to produce this input file.

It therefore infers that the {barcode} wildcard somehow has to contain this .consensus.fasta that it has never seen before. This wildcard is then handed over to the top, where the workflow crashes since it cannot find a matching input file.

Second: This initialisation of the wildcard with sth. you don't want is only possible since you did not constrain the wildcard properly. You can for example forbid the wildcard to contain a . (see wildcard_constraints here)

However, the main problem is that catFasta does not find the desired input. I'd suggest changing the output of minion to "nanopolish/{barcode}/{barcode}.consensus.fasta", since the you already take the OUTDIR from the params, that should not hurt your rule here.

Edit: Dummy test example:

(I think) The problem lies somewhere else in your code.

You have defined two rules, the first that outputs directory assembled, the second that outputs assembled/assembly.fasta. Since the output of the second rule is always at least the directory assembled, Snakemake complains. You can solve it by using the directory as input: