mgatk lite: Mitochondrial Genome Analysis Toolkit

mgatk-lite is a comprehensive toolkit designed for the analysis of mitochondrial genomes, originally developed by Caleb Lareau. The original provides a suite of tools to process and analyze mitochondrial DNA (mtDNA) from single-cell sequencing data, particularly from 10x Genomics platforms.

This version, however, has now removed most flexible functionality, with the purpose of optimizing and streamlining the tenx mode to work with 10x Genomics scATAC/ASAP-seq data.

Some of the options may not work as in our lab we generally run the program as:

mgatk-lite \
    -i ${project_outs}/${library}/outs/possorted_bam.bam \
    -n output \
    -o ${project_outs}/${library}/mgatk \
    -c $(nproc) \
    -bt CB \
    -b ${project_outs}/${library}/outs/filtered_peak_bc_matrix/barcodes.tsv \
    --skip-R \
    --remove-snakemake

Features

• Single-cell mtDNA Analysis: Process and analyze mtDNA from single-cell ATAC-seq
• Variant Calling: Identify and quantify mtDNA variants across single cells.
• Quality Control: Perform extensive quality control on mtDNA sequencing data.

Installation

To install mgatk-lite, use the following command:

conda create -y -n mgatk openjdk r-base r-data.table r-matrix bioconductor-genomicranges bioconductor-summarizedexperiment 
pip install pulp==2.7.0 matplotlib
git clone https://github.com/ollieeknight/mgatk-lite
cd mgatk-lite
pip install .

Test usage

git clone https://github.com/ollieeknight/mgatk-lite
cd mgatk-lite/tests/

mgatk-lite -i barcode/test_barcode.bam -n bc1 -o bc1dmem -bt CB -b barcode/test_barcodes.txt -c 2

2024-12-03 11:08:47,969 - INFO - mgatk version 1.0.0
2024-12-03 11:08:48,307 - INFO - Configuration for analysis:
2024-12-03 11:08:48,307 - INFO - Mode                      : tenx
2024-12-03 11:08:48,307 - INFO - Input                     : barcode/test_barcode.bam
2024-12-03 11:08:48,307 - INFO - Output                    : bc1dmem
2024-12-03 11:08:48,307 - INFO - Name                      : bc1
2024-12-03 11:08:48,307 - INFO - Mitochondrial genome      : rCRS
2024-12-03 11:08:48,307 - INFO - Number of cores           : 2
2024-12-03 11:08:48,307 - INFO - Barcode tag               : CB
2024-12-03 11:08:48,308 - INFO - Barcodes                  : barcode/test_barcodes.txt
2024-12-03 11:08:48,308 - INFO - Min barcode reads         : 1000
2024-12-03 11:08:48,308 - INFO - NHmax                     : 1
2024-12-03 11:08:48,308 - INFO - NMmax                     : 4
2024-12-03 11:08:48,308 - INFO - Remove duplicates         : False
2024-12-03 11:08:48,308 - INFO - UMI barcode               : XX
2024-12-03 11:08:48,308 - INFO - Handle overlap            : False
2024-12-03 11:08:48,308 - INFO - Low coverage threshold    : 10
2024-12-03 11:08:48,308 - INFO - Max java memory           : 8000m
2024-12-03 11:08:48,308 - INFO - Proper pairs              : False
2024-12-03 11:08:48,308 - INFO - Base quality              : 0
2024-12-03 11:08:48,308 - INFO - Alignment quality         : 0
2024-12-03 11:08:48,308 - INFO - Emit base qualities       : False
2024-12-03 11:08:48,308 - INFO - Number of samples         : 0
2024-12-03 11:08:48,308 - INFO - Keep samples              : ALL
2024-12-03 11:08:48,308 - INFO - Ignore samples            : NONE
2024-12-03 11:08:48,308 - INFO - Keep temp files           : False
2024-12-03 11:08:48,308 - INFO - Keep QC .bam files        : False
2024-12-03 11:08:48,308 - INFO - Skip R                    : False
2024-12-03 11:08:48,308 - INFO - Snake stdout              : False
2024-12-03 11:08:48,308 - INFO - Remove .snakemake         : False
2024-12-03 11:08:48,330 - INFO - Using fasta file: bc1dmem/fasta/chrM.fasta
2024-12-03 11:08:50,618 - INFO - Executing Snakemake command: snakemake --snakefile /home/olive/bin/mgatk-lite/mgatk/bin/snake/Snakefile.tenx --cores 2 --config cfp="bc1dmem/.internal/parseltongue/snake.scatter.yaml" > bc1dmem/logs/snakemake.log 2>&1
2024-12-03 11:09:22,163 - INFO - Successfully created final output files. Analysis complete.

Options

• --input, -i: Input .bam file from 10x single cell ATAC library.
• --output, -o: Output directory for analysis. Default is mgatk.
• --name, -n: Prefix for project name. Default is mgatk.
• --mito-genome, -g: Mitochondrial genome configuration. Default is rCRS.
• --ncores, -c: Number of cores to run the main job in parallel. Default is 1.
• --barcode-tag, -bt: Read tag to separate single cells.
• --barcodes, -b: Path to a file containing known barcodes.
• --min-barcode-reads, -mb: Minimum number of mitochondrial reads for a barcode to be genotyped. Default is 1000.
• --NHmax: Maximum number of read alignments allowed. Default is 1.
• --NMmax: Maximum number of paired mismatches allowed. Default is 4.
• --remove-duplicates, -rd: Remove duplicate reads.
• --umi-barcode, -ub: Read tag to specify the UMI tag when removing duplicates.
• --handle-overlap, -ho: Only count each base in the overlap region between a pair of reads once.
• --low-coverage-threshold, -lc: Variant count for each cell will be ignored below this threshold. Default is 10.
• --max-javamem, -jm: Maximum memory for Java for running duplicate removal per core. Default is 8000m.
• --proper-pairs, -pp: Require reads to be properly paired.
• --base-qual, -q: Minimum base quality for inclusion in the genotype count. Default is 0.
• --alignment-quality, -aq: Minimum alignment quality to include read in genotype. Default is 0.
• --emit-base-qualities, -eb: Output mean base quality per alt allele as part of the final output.
• --nsamples, -ns: Number of samples/cells to be processed per iteration. Default is 0 (all).
• --keep-samples, -k: Comma-separated list of sample names to keep. Default is ALL.
• --ignore-samples, -x: Comma-separated list of sample names to ignore. Default is NONE.
• --keep-temp-files, -z: Keep all intermediate files.
• --keep-qc-bams, -qc: Keep the quality-controlled bams after processing.
• --skip-R, -sr: Generate plain-text only output.
• --snake-stdout, -so: Write Snakemake log to stdout.
• --remove-snakemake, -rs: Delete the .snakemake directory once successfully run.

For more details, visit the mgatk GitHub repository.

Contact

The mgatk-lite package was developed and is maintained by Caleb Lareau, but this repo is maintained by Oliver Knight.