Snakemake - parameter file treated as a wildcard - bioinformatics

Question

I have written a pipeline in Snakemake. It's an ATAC-seq pipeline (bioinformatics pipeline to analyze genomics data from a specific experiment). Basically, until merging alignment step I use {sample_id} wildcard, to later switch to {sample} wildcard (merging two or more sample_ids into one sample).
working DAG here (for simplicity only one sample shown; orange and blue {sample_id}s are merged into one green {sample}
Tha all rule looks as follows:
configfile: "config.yaml"
SAMPLES_DICT = dict()
with open(config['SAMPLE_SHEET'], "r+") as fil:
next(fil)
for lin in fil.readlines():
row = lin.strip("\n").split("\t")
sample_id = row[0]
sample_name = row[1]
if sample_name in SAMPLES_DICT.keys():
SAMPLES_DICT[sample_name].append(sample_id)
else:
SAMPLES_DICT[sample_name] = [sample_id]
SAMPLES = list(SAMPLES_DICT.keys())
SAMPLE_IDS = [sample_id for sample in SAMPLES_DICT.values() for sample_id in sample]
rule all:
input:
# FASTQC output for RAW reads
expand(os.path.join(config['FASTQC'], '{sample_id}_R{read}_fastqc.zip'),
sample_id = SAMPLE_IDS,
read = ['1', '2']),
# Trimming
expand(os.path.join(config['TRIMMED'],
'{sample_id}_R{read}_val_{read}.fq.gz'),
sample_id = SAMPLE_IDS,
read = ['1', '2']),
# Alignment
expand(os.path.join(config['ALIGNMENT'], '{sample_id}_sorted.bam'),
sample_id = SAMPLE_IDS),
# Merging
expand(os.path.join(config['ALIGNMENT'], '{sample}_sorted_merged.bam'),
sample = SAMPLES),
# Marking Duplicates
expand(os.path.join(config['ALIGNMENT'], '{sample}_sorted_md.bam'),
sample = SAMPLES),
# Filtering
expand(os.path.join(config['FILTERED'],
'{sample}.bam'),
sample = SAMPLES),
expand(os.path.join(config['FILTERED'],
'{sample}.bam.bai'),
sample = SAMPLES),
# multiqc report
"multiqc_report.html"
message:
'\n#################### ATAC-seq pipeline #####################\n'
'Running all necessary rules to produce complete output.\n'
'############################################################'
I know it's too messy, I should only leave the necessary bits, but here my understanding of snakemake fails cause I don't know what I have to keep and what I should delete.
This is working, to my knowledge exactly as I want.
However, I added a rule:
rule hmmratac:
input:
bam = os.path.join(config['FILTERED'], '{sample}.bam'),
index = os.path.join(config['FILTERED'], '{sample}.bam.bai')
output:
model = os.path.join(config['HMMRATAC'], '{sample}.model'),
gappedPeak = os.path.join(config['HMMRATAC'], '{sample}_peaks.gappedPeak'),
summits = os.path.join(config['HMMRATAC'], '{sample}_summits.bed'),
states = os.path.join(config['HMMRATAC'], '{sample}.bedgraph'),
logs = os.path.join(config['HMMRATAC'], '{sample}.log'),
sample_name = '{sample}'
log:
os.path.join(config['LOGS'], 'hmmratac', '{sample}.log')
params:
genomes = config['GENOMES'],
blacklisted = config['BLACKLIST']
resources:
mem_mb = 32000
message:
'\n######################### Peak calling ########################\n'
'Peak calling for {output.sample_name}\n.'
'############################################################'
shell:
'HMMRATAC -Xms2g -Xmx{resources.mem_mb}m '
'--bam {input.bam} --index {input.index} '
'--genome {params.genome} --blacklist {params.blacklisted} '
'--output {output.sample_name} --bedgraph true &> {log}'
And into the rule all, after filtering, before multiqc, I added:
# Peak calling
expand(os.path.join(config['HMMRATAC'], '{sample}.model'),
sample = SAMPLES),
Relevant config.yaml fragments:
# Path to blacklisted regions
BLACKLIST: "/mnt/data/.../hg38.blacklist.bed"
# Path to chromosome sizes
GENOMES: "/mnt/data/.../hg38_sizes.genome"
# Path to filtered alignment
FILTERED: "alignment/filtered"
# Path to peaks
HMMRATAC: "peaks/hmmratac"
This is the error* I get (It goes on for every input and output of the rule). *Technically it's a warning but it halts execution of snakemake so I am calling it an error.
File path alignment/filtered//mnt/data/.../hg38.blacklist.bed.bam contains double '/'. This is likely unintended. It can also lead to inconsistent results of the file-matching approach used by Snakemake.
WARNING:snakemake.logging:File path alignment/filtered//mnt/data/.../hg38.blacklist.bed.bam contains double '/'. This is likely unintended. It can also lead to inconsistent results of the file-matching approach used by Snakemake.
It isn't actually ... - I just didn't feel safe providing an absolute path here.
For a couple of days, I have struggled with this error. Looked through the documentation, listened to the introduction. I understand that the above description is far from perfect (it is huge bc I don't even know how to work it down to provide minimal reproducible example...) but I am desperate and hope you can be patient with me.
Any suggestions as to how to google it, where to look for an error would be much appreciated.

Answer 1

Technically it's a warning but it halts execution of snakemake so I am calling it an error.
It would be useful to post the logs from snakemake to see if snakemake terminated with an error and if so what error.
However, in addition to Eric C.'s suggestion to use wildcards.sample instead of {sample} as file name, I think that this is quite suspicious:
alignment/filtered//mnt/data/.../hg38.blacklist.bed.bam
/mnt/ is usually at the root of the file system and you are prepending to it a relative path (alignment/filtered). Are you sure it is correct?