Snakefile

"""Hevesi et al., 2023"""
import pandas as pd
from os import listdir, rename, getcwd
from os.path import join, basename, dirname, abspath
from pathlib import Path
from snakemake.utils import validate, min_version

##### set minimum snakemake version #####
min_version("7.20.0")

##### load config and sample sheets #####
configfile: "config.yaml"
samples = pd.read_table(config["samples"]).set_index("cellranger", drop=False)

container: "docker://continuumio/miniconda3:4.4.10"

##### target rules #####

shell.executable("/bin/bash")

rule all:
    input:
        expand("souporcell/{cellranger}/clusters.tsv",
                cellranger=samples["cellranger"]),
        expand("cellbender/{cellranger}/{cellranger}_output.h5",
                cellranger=samples["cellranger"])

##### load rules #####

CELLRANGER="source /home/etretiakov/src/cellranger-7.1.0/sourceme.bash && cellranger "

rule cellranger_count:
    input:
        sample=directory("fastq/{cellranger}"),
        idx=directory("mm10_optimized")
    output:
        summary="cellranger/{cellranger}/outs/web_summary.html",
        bam="cellranger/{cellranger}/outs/possorted_genome_bam.bam",
    params:
        sample="cellranger/{cellranger}"
    threads: 32
    shell:
        ("{CELLRANGER} count --include-introns true \
            --id={params.sample} \
            --transcriptome={input.idx} \
            --fastqs={input.sample} \
            --jobmode=local \
            --localcores={threads} ")

rule souporcell:
    input:
        bam="cellranger/{cellranger}/outs/possorted_genome_bam.bam",
        barcodes="cellranger/{cellranger}/outs/filtered_feature_bc_matrix/barcodes.tsv.gz",
        fasta="mm10_optimized/fasta/genome.fa"
    output:
        dir=directory("souporcell/{cellranger}"),
        clusters="souporcell/{cellranger}/clusters.tsv"
    params: 2
    container:
        "shub://wheaton5/souporcell"
    threads: 20
    shell:
        ("souporcell_pipeline.py \
            -i {input.bam} \
            -b {input.barcodes} \
            -f {input.fasta} \
            -t {threads} \
            -o {output.dir} \
            -k {params}")

rule cellbender:
    input:
        "cellranger/{cellranger}/outs/raw_feature_bc_matrix.h5"
    output:
        "cellbender/{cellranger}/{cellranger}_output.h5"
    params:
        ndroplets=lambda wildcards: samples["ndroplets"][wildcards.cellranger],
        ncells=lambda wildcards: samples["ncells"][wildcards.cellranger]
    container:
        "docker://etretiakov/cellbender:v0.0.1"
    threads: 20
    resources:
        nvidia_gpu=1
    shell:
        ("cellbender remove-background \
            --input {input} \
            --output {output} \
            --cuda \
            --expected-cells {params.ncells} \
            --total-droplets-included {params.ndroplets} \
            --fpr 0.01 \
            --epochs 150")