SnakePolyester (v0.1.0)

A snakemake workflow for simulating RNA-seq reads by Polyester. You can generate RNA-seq reads for multiple scenarios with different coverage, replicates, and size factors.

This workflow generates RNA-seq read files in fastq format and maps them to the reference genome using STAR.

Install

• Make sure you have Singularity and Snakemake installed.
• Clone this repository.

git clone https://github.com/NaotoKubota/SnakePolyester.git

Usage

snakemake -s /path/to/SnakePolyester/SnakePolyester.smk --configfile config.yaml --cores 32 --use-singularity --singularity-args "--bind $HOME:$HOME" --rerun-incomplete

Input files

• transcript_table.tsv: A table containing transcript IDs and their fold-changes. Columns are transcript_id, fold-change in condition1, fold-change in condition2.

ENSMUST00000000003	1.0	1.4
ENSMUST00000000028	1.0	0.6
ENSMUST00000000080	1.0	1.0
ENSMUST00000000090	1.0	1.4
ENSMUST00000000153	1.0	1.0
ENSMUST00000000161	1.0	0.6
ENSMUST00000000284	1.0	1.4
ENSMUST00000000299	1.0	0.6
ENSMUST00000000304	1.0	1.0
ENSMUST00000000335	1.0	0.6

• config.yaml: Configuration file.

workdir:
    /path/to/outdir
transcript_table:
    /path/to/transcript_table.tsv
cdna_fasta:
    /path/to/Mus_musculus.GRCm38.cdna.all.fa # cDNA fasta file
star_index:
    /path/to/star_index # STAR index directory

scenarios:
    scenario1: # Scenario name
        coverage: # Coverage of the simulated reads for each transcript
            30
        replicates: # Number of replicates for each condition
            1
        size_factor: # controls the per-transcript mean/variance relationship. Increase the value of size factor to introduce more variance into your simulations.
            3 # low-variance situation
    scenario2:
        coverage:
            30
        replicates:
            2
        size_factor:
            3
    scenario3:
        coverage:
            30
        replicates:
            3
        size_factor:
            3

Output files

• benchmark: Benchmarking results.
• data: Simulated RNA-seq read files.
• logs: Log files.

outdir/
├── benchmark
├── data
└── logs

An example of data directory structure

outdir/data/
├── scenario1
│   ├── bam_table.tsv
│   ├── fastq_table.tsv
│   ├── flag.txt
│   ├── polyester
│   │   ├── sample_01_1.fastq.gz
│   │   ├── sample_01_2.fastq.gz
│   │   ├── sample_02_1.fastq.gz
│   │   ├── sample_02_2.fastq.gz
│   │   ├── sim_counts_matrix.rda
│   │   ├── sim_rep_info.txt
│   │   └── sim_tx_info.txt
│   └── star
│       ├── sample_01
│       │   ├── sample_01_Aligned.out.bam
│       │   ├── sample_01_Aligned.out.bam.bai
│       │   ├── sample_01_Log.final.out
│       │   ├── sample_01_Log.out
│       │   ├── sample_01_Log.progress.out
│       │   └── sample_01_SJ.out.tab
│       └── sample_02
│           ├── sample_02_Aligned.out.bam
│           ├── sample_02_Aligned.out.bam.bai
│           ├── sample_02_Log.final.out
│           ├── sample_02_Log.out
│           ├── sample_02_Log.progress.out
│           └── sample_02_SJ.out.tab
├── scenario2
│   ├── bam_table.tsv
│   ├── fastq_table.tsv
│   ├── flag.txt
│   ├── polyester
│   │   ├── sample_01_1.fastq.gz
│   │   ├── sample_01_2.fastq.gz
│   │   ├── sample_02_1.fastq.gz
│   │   ├── sample_02_2.fastq.gz
│   │   ├── sample_03_1.fastq.gz
│   │   ├── sample_03_2.fastq.gz
│   │   ├── sample_04_1.fasta
│   │   ├── sample_04_2.fasta
│   │   ├── sim_counts_matrix.rda
│   │   ├── sim_rep_info.txt
│   │   └── sim_tx_info.txt
│   └── star
│       ├── sample_01
│       │   ├── sample_01_Aligned.out.bam
│       │   ├── sample_01_Aligned.out.bam.bai
│       │   ├── sample_01_Log.final.out
│       │   ├── sample_01_Log.out
│       │   ├── sample_01_Log.progress.out
│       │   └── sample_01_SJ.out.tab
│       ├── sample_02
│       │   ├── sample_02_Aligned.out.bam
│       │   ├── sample_02_Aligned.out.bam.bai
│       │   ├── sample_02_Log.final.out
│       │   ├── sample_02_Log.out
│       │   ├── sample_02_Log.progress.out
│       │   └── sample_02_SJ.out.tab
│       └── sample_03
│           ├── sample_03_Aligned.out.bam
│           ├── sample_03_Aligned.out.bam.bai
│           ├── sample_03_Log.final.out
│           ├── sample_03_Log.out
│           ├── sample_03_Log.progress.out
│           └── sample_03_SJ.out.tab
├── scenario3
│   ├── bam_table.tsv
│   ├── fastq_table.tsv
│   ├── flag.txt
│   ├── polyester
│   │   ├── sample_01_1.fastq.gz
│   │   ├── sample_01_2.fastq.gz
│   │   ├── sample_02_1.fastq.gz
│   │   ├── sample_02_2.fastq.gz
│   │   ├── sample_03_1.fastq.gz
│   │   ├── sample_03_2.fastq.gz
│   │   ├── sample_04_1.fastq.gz
│   │   ├── sample_04_2.fastq.gz
│   │   ├── sample_05_1.fasta
│   │   ├── sample_05_2.fasta
│   │   ├── sample_06_1.fasta
│   │   ├── sample_06_2.fasta
│   │   ├── sim_counts_matrix.rda
│   │   ├── sim_rep_info.txt
│   │   └── sim_tx_info.txt
│   └── star
│       ├── sample_01
│       │   ├── sample_01_Aligned.out.bam
│       │   ├── sample_01_Aligned.out.bam.bai
│       │   ├── sample_01_Log.final.out
│       │   ├── sample_01_Log.out
│       │   ├── sample_01_Log.progress.out
│       │   └── sample_01_SJ.out.tab
│       ├── sample_02
│       │   ├── sample_02_Aligned.out.bam
│       │   ├── sample_02_Aligned.out.bam.bai
│       │   ├── sample_02_Log.final.out
│       │   ├── sample_02_Log.out
│       │   ├── sample_02_Log.progress.out
│       │   └── sample_02_SJ.out.tab
│       ├── sample_03
│       │   ├── sample_03_Aligned.out.bam
│       │   ├── sample_03_Aligned.out.bam.bai
│       │   ├── sample_03_Log.final.out
│       │   ├── sample_03_Log.out
│       │   ├── sample_03_Log.progress.out
│       │   └── sample_03_SJ.out.tab
│       └── sample_04
│           ├── sample_04_Aligned.out.bam
│           ├── sample_04_Aligned.out.bam.bai
│           ├── sample_04_Log.final.out
│           ├── sample_04_Log.out
│           ├── sample_04_Log.progress.out
│           └── sample_04_SJ.out.tab
└── transcript_cdna.fa