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'''
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'''
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config["output_dir"],
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"{sample}",
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reads2 = os.path.join(
config["output_dir"],
"samples",
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"{sample}",
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reads1 = temp(os.path.join(
config["output_dir"],
"{sample}.pe.remove_adapters_mate1.fastq.gz")),
reads2 = temp(os.path.join(
config["output_dir"],
"{sample}.pe.remove_adapters_mate2.fastq.gz"))
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params:
adapter_3_mate1 = lambda wildcards:
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get_sample('fq1_3p', search_id='index', search_value=wildcards.sample),
adapter_5_mate1 = lambda wildcards:
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get_sample('fq1_5p', search_id='index', search_value=wildcards.sample),
adapter_3_mate2 = lambda wildcards:
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get_sample('fq2_3p', search_id='index', search_value=wildcards.sample),
adapter_5_mate2 = lambda wildcards:
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get_sample('fq2_5p', search_id='index', search_value=wildcards.sample)
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singularity:
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stderr = os.path.join(
config["log_dir"],
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"{sample}",
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stdout = os.path.join(
config["log_dir"],
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"{sample}",
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shell:
"(cutadapt \
-j {threads} \
-m 10 \
-a {params.adapter_3_mate1} \
-g {params.adapter_5_mate1} \
-A {params.adapter_3_mate2} \
-G {params.adapter_5_mate2} \
-o {output.reads1} \
-p {output.reads2} \
{input.reads1} \
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1> {log.stdout} 2>{log.stderr}"
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rule pe_remove_polya_cutadapt:
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'''
Remove polyA tails
'''
input:
reads1 = os.path.join(
config["output_dir"],
"{sample}.pe.remove_adapters_mate1.fastq.gz"),
reads2 = os.path.join(
config["output_dir"],
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reads1 = temp(os.path.join(
config["output_dir"],
"{sample}.pe.remove_polya_mate1.fastq.gz")),
reads2 = temp(os.path.join(
config["output_dir"],
"{sample}",
"{sample}.pe.remove_polya_mate2.fastq.gz"))
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params:
polya_3_mate1 = lambda wildcards:
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get_sample(
'fq1_polya_3p',
search_id='index',
search_value=wildcards.sample),
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get_sample(
'fq1_polya_5p',
search_id='index',
search_value=wildcards.sample),
polya_3_mate2 = lambda wildcards:
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get_sample(
'fq2_polya_3p',
search_id='index',
search_value=wildcards.sample),
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get_sample(
'fq2_polya_5p',
search_id='index',
search_value=wildcards.sample)
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singularity:
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stderr = os.path.join(
config["log_dir"],
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"{sample}",
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stdout = os.path.join(
config["log_dir"],
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"{sample}",
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"(cutadapt \
-j {threads} \
-m 10 \
-a {params.polya_3_mate1} \
-A {params.polya_3_mate2} \
-o {output.reads1} \
-p {output.reads2} \
{input.reads1} \
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1> {log.stdout} 2>{log.stderr}"
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rule pe_map_genome_star:
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'''
Map to genome using STAR
'''
input:
index = lambda wildcards:
os.path.join(
config["star_indexes"],
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get_sample(
'organism',
search_id='index',
search_value=wildcards.sample),
get_sample(
'index_size',
search_id='index',
search_value=wildcards.sample),
"STAR_index",
"chrNameLength.txt"),
reads1 = os.path.join(
config["output_dir"],
"{sample}",
reads2 = os.path.join(
config["output_dir"],
"{sample}",
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output:
bam = os.path.join(
config["output_dir"],
"{sample}",
"map_genome",
"{sample}.pe.Aligned.sortedByCoord.out.bam"),
logfile = os.path.join(
config["output_dir"],
"{sample}",
"map_genome",
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params:
sample_id = "{sample}",
index = lambda wildcards:
os.path.abspath(os.path.join(
config["star_indexes"],
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get_sample(
'organism',
search_id='index',
search_value=wildcards.sample),
get_sample(
'index_size',
search_id='index',
search_value=wildcards.sample),
outFileNamePrefix = os.path.join(
config["output_dir"],
"{sample}",
"map_genome",
multimappers = lambda wildcards:
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get_sample(
'multimappers',
search_id='index',
search_value=wildcards.sample),
soft_clip = lambda wildcards:
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get_sample(
'soft_clip',
search_id='index',
search_value=wildcards.sample),
pass_mode = lambda wildcards:
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get_sample(
'pass_mode',
search_id='index',
search_value=wildcards.sample),
singularity:
threads: 12
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stderr = os.path.join(
config["log_dir"],
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"{sample}",
shell:
"(STAR \
--twopassMode {params.pass_mode} \
--runThreadN {threads} \
--genomeDir {params.index} \
--readFilesIn {input.reads1} {input.reads2} \
--readFilesCommand zcat \
--outFilterMultimapNmax {params.multimappers} \
--outFileNamePrefix {params.outFileNamePrefix} \
--outSAMattributes All \
--outStd BAM_SortedByCoordinate \
--outSAMtype BAM SortedByCoordinate \
--outFilterType BySJout \
--outSAMattrRGline ID:rnaseq_pipeline SM:{params.sample_id} \
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--alignEndsType {params.soft_clip} > {output.bam};) \
2> {log.stderr}"
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rule pe_quantification_salmon:
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'''
Quantification at transcript and gene level using Salmon
'''
input:
reads1 = os.path.join(
config["output_dir"],
"{sample}",
reads2 = os.path.join(
config["output_dir"],
"{sample}",
gtf = lambda wildcards:
os.path.abspath(get_sample(
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'gtf',
search_id='index',
search_value=wildcards.sample)),
index = lambda wildcards:
os.path.join(
config["salmon_indexes"],
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get_sample(
'organism',
search_id='index',
search_value=wildcards.sample),
get_sample(
'kmer',
search_id='index',
search_value=wildcards.sample),
"salmon.idx")
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output:
gn_estimates = os.path.join(
config["output_dir"],
"{sample}",
"quant.genes.sf"),
tr_estimates = os.path.join(
config["output_dir"],
"{sample}",
"quant.sf"),
meta_info = os.path.join(
config["output_dir"],
"samples",
"{sample}",
"{sample}.salmon.pe",
"aux_info",
"meta_info.json"),
flenDist = os.path.join(
config["output_dir"],
"samples",
"{sample}",
"{sample}.salmon.pe",
"libParams",
"flenDist.txt")
shadow: "minimal"
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params:
output_dir = os.path.join(
config["output_dir"],
"{sample}",
libType = lambda wildcards:
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get_sample(
'libtype',
search_id='index',
search_value=wildcards.sample)
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stderr = os.path.join(
config["log_dir"],
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"{sample}",
"genome_quantification_salmon.pe.stderr.log"),
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stdout = os.path.join(
config["log_dir"],
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"{sample}",
"genome_quantification_salmon.pe.stdout.log"),
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threads: 6
singularity:
"docker://zavolab/salmon:1.1.0-slim"
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shell:
"(salmon quant \
--libType {params.libType} \
--seqBias \
--validateMappings \
--threads {threads} \
--writeUnmappedNames \
--index {input.index} \
--geneMap {input.gtf} \
-1 {input.reads1} \
-2 {input.reads2} \
-o {params.output_dir}; \
) 1> {log.stdout} 2> {log.stderr}"
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rule pe_genome_quantification_kallisto:
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'''
Quantification at transcript and gene level using Kallisto
'''
input:
reads1 = os.path.join(
config["output_dir"],
"{sample}",
reads2 = os.path.join(
config["output_dir"],
"{sample}",
index = lambda wildcards:
os.path.join(
config["kallisto_indexes"],
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get_sample(
'organism',
search_id='index',
search_value=wildcards.sample),
"kallisto.idx")
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output:
pseudoalignment = os.path.join(
config["output_dir"],
"{sample}",
"quant_kallisto",
"{sample}.pe.kallisto.pseudo.sam"),
abundances = os.path.join(
config["output_dir"],
"samples",
"{sample}",
"quant_kallisto",
"abundance.h5")
shadow: "minimal"
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params:
output_dir = os.path.join(
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config["output_dir"],
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"{sample}",
"quant_kallisto"),
directionality = lambda wildcards:
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get_sample(
'kallisto_directionality',
search_id='index',
search_value=wildcards.sample)
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singularity:
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threads: 8
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stderr = os.path.join(
config["log_dir"],
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"{sample}",
"genome_quantification_kallisto.pe.stderr.log")
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shell:
"(kallisto quant \
-i {input.index} \
-o {params.output_dir} \
--pseudobam \
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{input.reads1} {input.reads2} > {output.pseudoalignment}) \