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Snakefile
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Snakefile
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import pandas as pd
import numpy as np
from snakemake.utils import validate, min_version
##### set minimum snakemake version #####
min_version("5.1.2")
##### load config and sample sheets #####
configfile: "config.yaml"
validate(config, schema="06_Schemas/config.schema.yaml")
#### Set variables ####
ROOTDIR = os.getcwd()
RAWDATA = srcdir("00_RawData/")
REF = srcdir("01_Reference/")
CONTAINER = srcdir("02_Container/")
SCRIPTDIR = srcdir("03_Script/")
ENVDIR = srcdir("04_Workflow/")
OUTPUTDIR = srcdir("05_Output/")
REPORT = srcdir("07_Report/")
BENCHMARK = srcdir("08_benchmark/")
LOG = srcdir("09_log/")
# If running the workflow with slurm
run_slurm = config["run_slurm"]
if run_slurm:
container: CONTAINER + "mambaforge:23.1.0-1.sif"
# ----------------------------------------------
# Load sample sheet
# ----------------------------------------------
rawsample = pd.read_table(config["sample"]).set_index(["rawsample"], drop=False)
sample_id = pd.read_table(config["sample"]).set_index(["id"], drop=False)
sample_name = pd.read_table(config["sample"]).set_index(["name"], drop=False)
RAWSAMPLE = expand("{rawsample.rawsample}", rawsample = rawsample.itertuples()) # 2_S1_L001, 3_S1_L001 ...
SAMPLE_ID = expand("{sample_id.id}", sample_id = sample_id.itertuples()) # P5_KO,P5_WT,P30_KO,P30_WT
SAMPLE_NAME = expand("{sample_name.name}", sample_name = sample_name.itertuples()) # 2,3,6,7
# ----------------------------------------------
# Target rules
# ----------------------------------------------
TYPES = config["run"]["types"].split(',')
LIBRARY = config["run"]["library"].split(',')
SAMPLE = config["fastq"]["sname"].split(',')
NPROJ = config["fastq"]["nproject"]
FEATURES = config["diffexp"]["features"].split(',')
CELLMARKER = config["diffexp"]["cell_marker"].split(',')
AGRR = config["fastq"]["nproject"] # => A voir car varibale identique à NPROJ
NUM = config["fastq"]["pair"].split(',')
CLUSTER = config["diffexpsubset"]["cluster"].split(',')
rule all:
input:
### fastqc ###
# fastqc_output = expand(OUTPUTDIR + "00_clean/fastqc_output.txt"),
### multiqc ###
# multiqc_output = expand(OUTPUTDIR + "00_clean/multiqc_output.txt"),
# raw_multiqc_html = expand(OUTPUTDIR + "00_clean/raw_multiqc.html"),
### ReferenceEnhancer to generate a scRNA-seq optimized transcriptomic reference ###
### To improve with manual curration of the file overlapping_gene_list
# reference_enhancer_output = expand(OUTPUTDIR + "01_cellranger/reference_enhancer_output.txt"),
### reference for cellranger ###
# ref_cellranger_output = expand(OUTPUTDIR + "01_cellranger/ref_cellranger_output.txt"),
### Cell Multiplexing with cellranger multi ###
# cellranger_output = expand(OUTPUTDIR + "01_cellranger/cellranger_output.txt"),
# cellranger_html = expand(OUTPUTDIR + "01_cellranger/{sample_id}/web_summary.html", sample_id = SAMPLE_ID),
## If you need to aggregate your data
# aggrcsv = ROOTDIR + "/aggregation.csv",
# out_aggregate = expand(OUTPUTDIR + "01_cellranger/{agrr}/outs/aggregate_web_summary.html", agrr=AGRR),
## If demultiplexing is used
# bcf = OUTPUTDIR + "01_cellranger/Mix_MM_lines/outs/demuxlet_Mix_MM_lines.bcf",
# demuxlet = OUTPUTDIR + "01_cellranger/Mix_MM_lines/outs/demuxlet_Mix_MM_lines.best",
# tabdemuxlet = OUTPUTDIR + "01_cellranger/Mix_MM_lines/outs/demuxlet_Mix_MM_lines.tsv",
### Seurat ###
# seurat_report = expand(OUTPUTDIR + "02_seurat/{sample_id}/{sample_id}_seurat_report.html", sample_id = SAMPLE_ID),
### Normalisation with other method than Seurat ###
# normalisation_output = expand(OUTPUTDIR + "02_seurat/normalisation_output.txt"),
# NE MARCHE PAS : Doublet detection
# doublets_output = expand(OUTPUTDIR + "rm_doublet/doublets_output.txt"),
### Prepare data for SIMS
# data_for_sims_output = expand(OUTPUTDIR + "03_sims/data_for_sims_output.txt"),
# anndata_for_sims_output = expand(OUTPUTDIR + "03_sims/anndata_for_sims_output.txt"),
### KNNOR
# knnor_output = expand(OUTPUTDIR +"03_sims/knnor_output.txt"),
### SIMS
# sims_training_output = expand(OUTPUTDIR + "03_sims/output_sims_training.txt"),
# sims_prediction_output = expand(OUTPUTDIR + "03_sims/output_sims_prediction.txt"),
# sims_prediction_report = expand(OUTPUTDIR + "03_sims/{sample_id}/{sample_id}_data_matrix_prediction.csv", sample_id = SAMPLE_ID),
# sims_prediction_unknown_report = expand(OUTPUTDIR + "03_sims/{sample_id}/{sample_id}_data_matrix_prediction_filtered.csv", sample_id = SAMPLE_ID),
unknown_prediction_output = expand(OUTPUTDIR + "03_sims/unknown_prediction_output.txt"),
evaluate_prediction_output = expand(OUTPUTDIR + "03_sims/evaluate_prediction_output.txt"),
### Representing cellular assignation on UMAP
# umapAssignation_output = expand(OUTPUTDIR + "03_sims/umapAssignation_output.txt"),
# umap_sims_report = expand(OUTPUTDIR + "03_sims/{sample_id}/{sample_id}_umap_sims.pdf", sample_id = SAMPLE_ID),
# umap_sims_threshold_report = expand(OUTPUTDIR + "03_sims/{sample_id}/{sample_id}_umap_sims_threshold.pdf", sample_id = SAMPLE_ID),
# umap_per_labels_report = expand(OUTPUTDIR + "03_sims/{sample_id}/{sample_id}_umap_per_labels.pdf", sample_id = SAMPLE_ID),
## Differential expression analyses
# violinplot = expand(OUTPUTDIR + "03_diffexp/violin_plot/{features}_violin_plot.pdf", features=FEATURES),
# umapfeature = expand(OUTPUTDIR + "03_diffexp/umap_plot/{features}_umapfeature_plot.pdf", features=FEATURES),
# tsnefeature = expand(OUTPUTDIR + "03_diffexp/tsne_plot/{features}_tsnefeature_plot.pdf", features=FEATURES),
# ridgefeature = expand(OUTPUTDIR + "03_diffexp/ridge_plot/{features}_ridgefeature_plot.pdf", features=FEATURES),
# heatmapfeature = OUTPUTDIR + "03_diffexp/heatmap/heatmapfeature.pdf",
# diffexp_report = OUTPUTDIR + "03_diffexp/" + NPROJ + "_diffexp_report.html",
# defile_allcells = expand(OUTPUTDIR + "03_diffexp/differential_expression_features/{wt}_vs_AllCellMarker_DE.csv", wt=WT),
# defile = expand(OUTPUTDIR + "03_diffexp/differential_expression_features/{wt}_vs_{cellmarker}_DE.csv", wt=WT, cellmarker=CELLMARKER),
# volcano_allcluster = expand(OUTPUTDIR + "03_diffexp/volcanoplot/volcano_{wt}_vs_AllCellMarker.pdf", wt=WT),
# volcano = expand(OUTPUTDIR + "03_diffexp/volcanoplot/volcano_{wt}_vs_{cellmarker}.pdf", wt=WT, cellmarker=CELLMARKER),
# sign_up_allcell = expand(OUTPUTDIR + "03_diffexp/de_significant/{wt}_vs_AllCellMarker_signif-up-regulated.txt", wt=WT),
# sign_up = expand(OUTPUTDIR + "03_diffexp/de_significant/{wt}_vs_{cellmarker}_signif-up-regulated.txt", wt=WT, cellmarker=CELLMARKER),
# sign_down_allcell = expand(OUTPUTDIR + "03_diffexp/de_significant/{wt}_vs_AllCellMarker_signif-down-regulated.txt", wt=WT),
# sign_down = expand(OUTPUTDIR + "03_diffexp/de_significant/{wt}_vs_{cellmarker}_signif-down-regulated.txt", wt=WT, cellmarker=CELLMARKER),
### Differential expression analyses inside each cluster after subset (by gene expression)
# diffexp_subset_report = OUTPUTDIR + "04_diffexp_subset/" + NPROJ + "_diffexp_subset_report.html",
# defile_subset = expand(OUTPUTDIR + "04_diffexp_subset/differential_expression/{cluster}_subcluster_DE.csv", cluster=CLUSTER),
# volcano_subset = expand(OUTPUTDIR + "04_diffexp_subset/volcanoplot/volcano_{cluster}.pdf", cluster=CLUSTER),
# sign_up_subset = expand(OUTPUTDIR + "04_diffexp_subset/de_significant/{cluster}_signif-up-regulated.txt", cluster=CLUSTER),
# sign_down_subset = expand(OUTPUTDIR + "04_diffexp_subset/de_significant/{cluster}_signif-down-regulated.txt", cluster=CLUSTER),
# violinplot_subset = expand(OUTPUTDIR + "04_diffexp_subset/violin_plot/{features}_violin_plot.pdf", features=FEATURES),
### Compress result files and figures ###
# data_matrix_tar = OUTPUTDIR + "02_seurat/data_matrix.tar.gz",
# violinplot_tar = OUTPUTDIR + "03_diffexp/violin_plot.tar.gz",
# umapfeature_tar = OUTPUTDIR + "03_diffexp/umapfeature_plot.tar.gz",
# tsnefeature_tar = OUTPUTDIR + "03_diffexp/tsnefeature_plot.tar.gz",
# ridgefeature_tar = OUTPUTDIR + "03_diffexp/ridgefeature_plot.tar.gz",
# heatmapfeature_tar = OUTPUTDIR + "03_diffexp/heatmapfeature.tar.gz",
# defile_tar = OUTPUTDIR + "03_diffexp/differencial_expression_tests.tar.gz",
# volcano_tar = OUTPUTDIR + "03_diffexp/volcano_plot.tar.gz",
# sign_up_down_tar = OUTPUTDIR + "03_diffexp/up_down_regulated_genes_list.tar.gz",
# defile_subset_tar = OUTPUTDIR + "04_diffexp_sub
# ----------------------------------------------
# setup report
# ----------------------------------------------
report: "07_Report/workflow.rst"
# ----------------------------------------------
# Load rules
# ----------------------------------------------
run_demultiplex = config["run_demultiplex"]
run_multiplex = config["run_multiplex"]
if run_demultiplex:
include: ENVDIR + "clean_demultiplex.smk"
include: ENVDIR + "cellranger_demultiplex.smk"
include: ENVDIR + "seurat.smk"
include: ENVDIR + "prepare_data_sims.smk"
include: ENVDIR + "SIMS.smk"
include: ENVDIR + "umapCellAssignation.smk"
include: ENVDIR + "knnor.smk"
if run_multiplex:
# include: ENVDIR + "clean.smk"
# include: ENVDIR + "cellranger.smk"
# NE MARCHE PAS include: ENVDIR + "processing_multiplex.smk"
# include: ENVDIR + "seurat.smk"
# NE MARCHE PAS include: ENVDIR + "rm_doublets.smk"
# include: ENVDIR + "prepare_data_sims.smk"
include: ENVDIR + "SIMS.smk"
# include: ENVDIR + "umapCellAssignation.smk"
# include: ENVDIR + "demuxlet.smk"
# include: ENVDIR + "seurat.smk"
# include: ENVDIR + "diffexp.smk"
# include: ENVDIR + "diffexp_subset.smk"
# include: ENVDIR + "report.smk"