common.smk 22.4 KB
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from snakemake.utils import validate
import pandas as pd
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import os
from smart_open import open
import yaml
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# this container defines the underlying OS for each job when using the workflow
# with --use-conda --use-singularity
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# container: "docker://continuumio/miniconda3"
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##### load config and sample sheets #####

validate(config, schema="../schemas/config.schema.yaml")

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samples = pd.read_csv(config["samples"], sep="\t", dtype = str).set_index("sample", drop=False)
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samples.index.names = ["sample_id"]
validate(samples, schema="../schemas/samples.schema.yaml")
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units = pd.read_csv(
    config["units"], dtype=str, sep="\t").set_index(["sample", "unit"], drop=False)
units.index.names = ["sample_id", "unit_id"]
units.index = units.index.set_levels(
    [i.astype(str) for i in units.index.levels])  # enforce str in index
validate(units, schema="../schemas/units.schema.yaml")

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build = config["resources"]["ref"]["build"]
chromosome = config["resources"]["ref"]["chromosome"]
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##### wildcard constraints #####

wildcard_constraints:
    sample = "|".join(samples.index),
    unit = "|".join(units["unit"])

####### helpers ###########

def is_single_end(sample, unit):
    """Determine whether unit is single-end."""
    fq2_present = pd.isnull(units.loc[(sample, unit), "fq2"])
    if isinstance(fq2_present, pd.core.series.Series):
        # if this is the case, get_fastqs cannot work properly
        raise ValueError(
            f"Multiple fq2 entries found for sample-unit combination {sample}-{unit}.\n"
            "This is most likely due to a faulty units.tsv file, e.g. "
            "a unit name is used twice for the same sample.\n"
            "Try checking your units.tsv for duplicates."
        )
    return fq2_present

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def has_only_sra_accession(sample, unit):
    return pd.isnull(units.loc[(sample, unit), "fq1"]) and pd.isnull(units.loc[(sample, unit), "fq2"]) \
           and not pd.isnull(units.loc[(sample, unit), "sra_accession"])

def is_sra_se(sample, unit):
    return has_only_sra_accession(sample, unit) and config["single_end"]

def is_sra_pe(sample, unit):
    return has_only_sra_accession(sample, unit) and not config["single_end"]

def get_se_pe_branches_input(wildcards):
    if config["single_end"]:
        return "results/bamtools_filtered/{sample}.sorted.bam".format(sample=wildcards.sample)
    else:
        return "results/orph_rm_pe/{sample}.sorted.bam".format(sample=wildcards.sample)

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def get_individual_fastq(wildcards):
    """Get individual raw FASTQ files from unit sheet, based on a read (end) wildcard"""
    if ( wildcards.read == "0" or wildcards.read == "1" ):
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        if is_sra_se(wildcards.sample, wildcards.unit):
            return expand("resources/ref/sra-se-reads/{accession}.fastq",
                              accession=units.loc[ (wildcards.sample, wildcards.unit), "sra_accession" ])
        elif is_sra_pe(wildcards.sample, wildcards.unit):
            return expand("resources/ref/sra-pe-reads/{accession}.1.fastq",
                              accession=units.loc[ (wildcards.sample, wildcards.unit), "sra_accession" ])
        else:
            return units.loc[ (wildcards.sample, wildcards.unit), "fq1" ]
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    elif wildcards.read == "2":
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        if is_sra_pe(wildcards.sample, wildcards.unit):
            return expand("resources/ref/sra-pe-reads/{accession}.2.fastq",
                          accession=units.loc[ (wildcards.sample, wildcards.unit), "sra_accession" ])
        else:
            return units.loc[ (wildcards.sample, wildcards.unit), "fq2" ]
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def get_fastqs(wildcards):
    """Get raw FASTQ files from unit sheet."""
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    if is_sra_se(wildcards.sample, wildcards.unit):
        return expand("resources/ref/sra-se-reads/{accession}.fastq",
                          accession=units.loc[ (wildcards.sample, wildcards.unit), "sra_accession" ])
    elif is_sra_pe(wildcards.sample, wildcards.unit):
        return expand(["resources/ref/sra-pe-reads/{accession}.1.fastq", "ref/sra-pe-reads/{accession}.2.fastq"],
                          accession=units.loc[ (wildcards.sample, wildcards.unit), "sra_accession" ])
    elif is_single_end(wildcards.sample, wildcards.unit):
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        return units.loc[ (wildcards.sample, wildcards.unit), "fq1" ]
    else:
        u = units.loc[ (wildcards.sample, wildcards.unit), ["fq1", "fq2"] ].dropna()
        return [ f"{u.fq1}", f"{u.fq2}" ]

def is_control(sample):
    control = samples.loc[sample]["control"]
    return pd.isna(control) or pd.isnull(control)

def get_sample_control_peak_combinations_list():
    sam_contr = []
    for sample in samples.index:
        if not is_control(sample):
            sam_contr.extend(expand(["{sample}-{control}.{peak}"], sample = sample, control = samples.loc[sample]["control"], peak = config["params"]["peak-analysis"]))
    return sam_contr

def get_peaks_count_plot_input():
    return expand(
        "results/macs2_callpeak/peaks_count/{sam_contr_peak}.peaks_count.tsv",
        sam_contr_peak = get_sample_control_peak_combinations_list()
    )

def get_frip_score_input():
    return expand(
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        "results/bedtools_intersect/{sam_contr_peak}.peaks_frip.tsv",
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        sam_contr_peak = get_sample_control_peak_combinations_list()
    )

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def get_macs2_peaks():
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    return expand(
        "results/macs2_callpeak/{sam_contr_peak}_peaks.{peak}Peak",
        sam_contr_peak = get_sample_control_peak_combinations_list(), peak =config["params"]["peak-analysis"]
    )

def get_plot_homer_annotatepeaks_input():
    return expand("results/homer/annotate_peaks/{sam_contr_peak}_peaks.annotatePeaks.txt",
        sam_contr_peak = get_sample_control_peak_combinations_list()
    )

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def get_samtools_view_filter_input(wildcards):
    return ["results/picard_dedup/{sample}.bam", "resources/ref/blacklist.sorted.complement".format(
        prefix="chr{chr}_".format(chr=chromosome) if chromosome else "",
        build=build
    )]

def exists_multiple_groups(antibody):
    return len(samples[samples["antibody"] == antibody]["group"].unique()) > 1

def exists_replicates(antibody):
    return len(samples[samples["antibody"] == antibody]["sample"].unique()) > 1

def get_controls_of_antibody(antibody):
    groups = samples[samples["antibody"] == antibody]["group"]
    controls = samples[pd.isnull(samples["control"])]
    return controls[controls["group"].isin(list(groups))]["sample"]

def get_samples_of_antibody(antibody):
    return samples[samples["antibody"] == antibody]["sample"]

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def get_map_reads_input(wildcards):
    if is_single_end(wildcards.sample, wildcards.unit):
        return "results/trimmed/{sample}-{unit}.fastq.gz"
    return ["results/trimmed/{sample}-{unit}.1.fastq.gz", "results/trimmed/{sample}-{unit}.2.fastq.gz"]

def get_read_group(wildcards):
    """Denote sample name and platform in read group."""
    return r"-R '@RG\tID:{sample}-{unit}\tSM:{sample}-{unit}\tPL:{platform}'".format(
        sample=wildcards.sample,
        unit=wildcards.unit,
        platform=units.loc[(wildcards.sample, wildcards.unit), "platform"])

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# def get_deseq2_igv_input():
#     return [file for file in os.listdir("results/deseq2/FDR/bed_files/FDR_0.05_{antibody}.consensus_{peak}-peaks")]

def get_igv_input():
    igv_input = []
    igv_input.extend([
        "results/IGV/big_wig/merged_library.bigWig.igv.txt"
    ])
    igv_input.extend(
        expand(
            [
                "results/IGV/macs2_callpeak-{peak}/merged_library.{sample_control_peak}_peaks.igv.txt",

            ],
            sample_control_peak=get_sample_control_peak_combinations_list(),
            peak=config["params"]["peak-analysis"]
        )
    )
    unique_antibodies = set([antibody for antibody in samples["antibody"] if not pd.isnull(antibody)])
    for antibody in unique_antibodies:
        igv_input.extend(
            expand(
                [
                    "results/IGV/consensus/merged_library.{antibody}.consensus_{peak}-peaks.igv.txt",
                    "results/IGV/consensus/merged_library.{antibody}.consensus_{peak}-peaks.deseq2.FDR_0.05.igv.txt"
                ],
                antibody=antibody,
                peak=config["params"]["peak-analysis"]
            )
        )
    return igv_input

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def get_multiqc_input(wildcards):
    multiqc_input = []
    for (sample, unit) in units.index:
        reads = [ "1", "2" ]
        if is_single_end(sample, unit):
            reads = [ "0" ]
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            multiqc_input.extend(expand (["logs/cutadapt/{sample}-{unit}.se.log"],
            sample = sample, unit = unit))
        else:
            multiqc_input.extend(expand (["logs/cutadapt/{sample}-{unit}.pe.log"],
            sample = sample, unit = unit))
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        multiqc_input.extend(
            expand (
                [
                    "results/qc/fastqc/{sample}.{unit}.{reads}_fastqc.zip",
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                    "results/qc/fastqc/{sample}.{unit}.{reads}.html",
                    "results/mapped/{sample}-{unit}.mapped.flagstat",
                    "results/mapped/{sample}-{unit}.mapped.idxstats",
                    "results/mapped/{sample}-{unit}.mapped.stats.txt"
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                ],
                sample = sample,
                unit = unit,
                reads = reads
            )
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        )
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    for sample in samples.index:
        multiqc_input.extend(
            expand (
                [
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                    "results/picard_dedup/{sample}.metrics.txt",
                    "results/picard_dedup/{sample}.picard_dedup.flagstat",
                    "results/picard_dedup/{sample}.picard_dedup.idxstats",
                    "results/picard_dedup/{sample}.picard_dedup.stats.txt",
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                    "results/bamtools_filtered/{sample}.sorted.bamtools_filtered.flagstat",
                    "results/bamtools_filtered/{sample}.sorted.bamtools_filtered.idxstats",
                    "results/bamtools_filtered/{sample}.sorted.bamtools_filtered.stats.txt",
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                    "results/phantompeakqualtools/{sample}.phantompeak.spp.out",
                    "results/phantompeakqualtools/{sample}.spp_correlation_mqc.tsv",
                    "results/phantompeakqualtools/{sample}.spp_nsc_mqc.tsv",
                    "results/phantompeakqualtools/{sample}.spp_rsc_mqc.tsv"
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                ],
                sample = sample
            )
        )
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        if config["params"]["deeptools-plots"]["activate"]:
            multiqc_input.extend(
                expand(
                    [
                        "results/deeptools/plot_profile_data.tab"
                    ],
                    sample=sample
                )
            )

        if not config["single_end"]:
            multiqc_input.extend(
                expand (
                    [
                        "results/orph_rm_pe/{sample}.sorted.orph_rm_pe.idxstats",
                        "results/orph_rm_pe/{sample}.sorted.orph_rm_pe.flagstat",
                        "results/orph_rm_pe/{sample}.sorted.orph_rm_pe.stats.txt"
                    ],
                    sample = sample
                )
            )

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        if not is_control(sample):
            multiqc_input.extend(
                expand (
                    [
                        "results/deeptools/{sample}-{control}.fingerprint_qcmetrics.txt",
                        "results/deeptools/{sample}-{control}.fingerprint_counts.txt",
                        "results/macs2_callpeak/{sample}-{control}.{peak}_peaks.xls"
                    ],
                sample = sample,
                control = samples.loc[sample]["control"],
                peak = config["params"]["peak-analysis"]
            )
        )
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        if config["params"]["lc_extrap"]["activate"]:
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                multiqc_input.extend( expand(["results/preseq/{sample}.lc_extrap"], sample = sample))
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        if config["params"]["picard_metrics"]["activate"]:
            if not config["single_end"]:
                multiqc_input.extend(
                    expand(
                        [
                            "results/qc/multiple_metrics/{sample}.insert_size_metrics",
                            "results/qc/multiple_metrics/{sample}.insert_size_histogram.pdf",
                        ],
                    sample = sample
                )
            )
            multiqc_input.extend(
                expand (
                    [                        
                        "results/qc/multiple_metrics/{sample}.alignment_summary_metrics",
                        "results/qc/multiple_metrics/{sample}.base_distribution_by_cycle_metrics",
                        "results/qc/multiple_metrics/{sample}.base_distribution_by_cycle.pdf",
                        "results/qc/multiple_metrics/{sample}.quality_by_cycle_metrics",
                        "results/qc/multiple_metrics/{sample}.quality_by_cycle.pdf",
                        "results/qc/multiple_metrics/{sample}.quality_distribution_metrics",
                        "results/qc/multiple_metrics/{sample}.quality_distribution.pdf"
                    ], 
                sample = sample
            )
        )
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    return multiqc_input
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def all_input(wildcards):
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    do_annot = config["params"]["peak-annotation-analysis"]["activate"]
    do_peak_qc = config["params"]["peak-qc"]["activate"]
    do_consensus_peak = config["params"]["consensus-peak-analysis"]["activate"]
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    wanted_input = []

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    # QC with fastQC and multiQC, igv session
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    wanted_input.extend([
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        "results/qc/multiqc/multiqc.html",
        "results/IGV/igv_session.xml"
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    ])
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    # trimming reads
    for (sample, unit) in units.index:
        if is_single_end(sample, unit):
            wanted_input.extend(expand(
                    [
                        "results/trimmed/{sample}-{unit}.fastq.gz",
                        "results/trimmed/{sample}-{unit}.se.qc.txt"
                    ],
                    sample = sample,
                    unit = unit
                )
            )
        else:
            wanted_input.extend(
                expand (
                    [
                        "results/trimmed/{sample}-{unit}.1.fastq.gz",
                        "results/trimmed/{sample}-{unit}.2.fastq.gz",
                        "results/trimmed/{sample}-{unit}.pe.qc.txt"
                    ],
                    sample = sample,
                    unit = unit
            )
        )

    # mapping, merging and filtering bam-files
    for sample in samples.index:
        wanted_input.extend(
            expand (
                [
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                    # "results/IGV/big_wig/merged_library.bigWig.igv.txt",
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                    "results/phantompeakqualtools/{sample}.phantompeak.pdf"
                ],
                sample = sample
            )
        )
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        if config["params"]["deeptools-plots"]["activate"]:
            wanted_input.extend(
                expand(
                    [
                        "results/deeptools/plot_profile.pdf",
                        "results/deeptools/heatmap.pdf",
                        "results/deeptools/heatmap_matrix.tab"
                    ],
                    sample=sample
                )
            )

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        if not is_control(sample):
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            with checkpoints.get_gsize.get().output[0].open() as f:
                # only produce the following files, if a gsize is specified
                if f.read().strip() != "":
                    if do_annot:
                        wanted_input.extend(
                            expand(
                                [
                                    "results/homer/annotate_peaks/{sample}-{control}.{peak}_peaks.annotatePeaks.txt"
                                ],
                                sample = sample,
                                control = samples.loc[sample]["control"],
                                peak = config["params"]["peak-analysis"]
                            )
                        )
                        if do_peak_qc:
                            wanted_input.extend(
                                expand(
                                    [
                                        "results/homer/plots/plot_{peak}_annotatepeaks_summary.txt",
                                        "results/homer/plots/plot_{peak}_annotatepeaks.pdf"
                                    ],
                                    peak = config["params"]["peak-analysis"]
                                )
                            )
                    if do_consensus_peak:
                        for antibody in samples["antibody"]:
                            if exists_multiple_groups(antibody) or exists_replicates(antibody):
                                wanted_input.extend(
                                    expand(
                                        [
                                            "results/macs2_merged_expand/{antibody}.consensus_{peak}-peaks.boolean.saf",
                                            "results/macs2_merged_expand/plots/{antibody}.consensus_{peak}-peaks.boolean.intersect.plot.pdf",
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                                            # "results/IGV/consensus/merged_library.{antibody}.consensus_{peak}-peaks.igv.txt"
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                                        ],
                                        peak = config["params"]["peak-analysis"],
                                        antibody = antibody
                                    )
                                )
                                if do_annot:
                                    wanted_input.extend(
                                        expand(
                                            [
                                                "results/homer/annotate_consensus_peaks/{antibody}.consensus_{peak}-peaks.annotatePeaks.txt",
                                                "results/homer/annotate_consensus_peaks/{antibody}.consensus_{peak}-peaks.boolean.annotatePeaks.txt",
                                                "results/feature_counts/{antibody}.consensus_{peak}-peaks.featureCounts",
                                                "results/feature_counts/{antibody}.consensus_{peak}-peaks.featureCounts.summary",
                                                "results/feature_counts/{antibody}.consensus_{peak}-peaks.featureCounts.jcounts",
                                                "results/deseq2/dss_rld/{antibody}.consensus_{peak}-peaks.dds.rld.RData",
                                                "results/deseq2/plots/{antibody}.consensus_{peak}-peaks.pca_plot.pdf",
                                                "results/deseq2/plots/{antibody}.consensus_{peak}-peaks.heatmap_plot.pdf",
                                                "results/deseq2/pca_vals/{antibody}.consensus_{peak}-peaks.pca.vals.txt",
                                                "results/deseq2/dists/{antibody}.consensus_{peak}-peaks.sample.dists.txt",
                                                "results/deseq2/sizeFactors/{antibody}.consensus_{peak}-peaks.sizeFactors.RData",
                                                "results/deseq2/sizeFactors/{antibody}.consensus_{peak}-peaks.sizeFactors.sizeFactor.txt",
                                                "results/deseq2/results/{antibody}.consensus_{peak}-peaks.deseq2_results.txt",
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                                                "results/deseq2/FDR/results/FDR_0.01_{antibody}.consensus_{peak}-peaks",
                                                "results/deseq2/FDR/results/FDR_0.05_{antibody}.consensus_{peak}-peaks",
                                                "results/deseq2/FDR/bed_files/FDR_0.01_{antibody}.consensus_{peak}-peaks",
                                                "results/deseq2/FDR/bed_files/FDR_0.05_{antibody}.consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/MA_plots/FDR_0.01_{antibody}consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/MA_plots/FDR_0.05_{antibody}consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/volcano_plots/FDR_0.01_{antibody}consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/volcano_plots/FDR_0.05_{antibody}consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/correlation_heatmaps_{antibody}consensus_{peak}-peaks",
                                                "results/deseq2/comparison_plots/scatter_plots_{antibody}consensus_{peak}-peaks"
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                                            ],
                                            peak = config["params"]["peak-analysis"],
                                            antibody = antibody
                                        )
                                    )
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            wanted_input.extend(
                expand(
                    [
                        "results/deeptools/{sample}-{control}.plot_fingerprint.pdf",
                        "results/macs2_callpeak/{sample}-{control}.{peak}_treat_pileup.bdg",
                        "results/macs2_callpeak/{sample}-{control}.{peak}_control_lambda.bdg",
                        "results/macs2_callpeak/{sample}-{control}.{peak}_peaks.{peak}Peak",
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                        # "results/IGV/macs2_callpeak-{peak}/merged_library.{sample}-{control}.{peak}_peaks.igv.txt",
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                        "results/macs2_callpeak/plots/plot_{peak}_peaks_count.pdf",
                        "results/macs2_callpeak/plots/plot_{peak}_peaks_frip_score.pdf",
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                        "results/macs2_callpeak/plots/plot_{peak}_peaks_macs2.pdf"
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                    ],
                    sample = sample,
                    control = samples.loc[sample]["control"],
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                    peak = config["params"]["peak-analysis"],
                    antibody = samples.loc[sample]["antibody"]
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                )
            )
            if config["params"]["peak-analysis"] == "broad":
                wanted_input.extend(
                    expand(
                        [
                            "results/macs2_callpeak/{sample}-{control}.{peak}_peaks.gappedPeak"
                        ],
                        sample = sample,
                        control = samples.loc[sample]["control"],
                        peak = config["params"]["peak-analysis"]
                    )
                )
            if config["params"]["peak-analysis"] == "narrow":
                wanted_input.extend(
                    expand(
                        [
                            "results/macs2_callpeak/{sample}-{control}.{peak}_summits.bed"
                        ],
                        sample = sample,
                        control = samples.loc[sample]["control"],
                        peak = config["params"]["peak-analysis"]
                    )
                )
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    return wanted_input