Scripts and data for reproducing MLA's evaluation

Software used

Running MLA

• MetaGraph (development commit bd39d9594e77f276e674033b52d9b26ec3a281e3, includes KMC)

Simulating reads

• ART Illumina (v2.5.8)
• pbsim3 (commit v3.0.0)
• PacBio ccs (v6.4.0)

Software for parsing read simulation outputs and alignment results

• bbmap (v38.86)
• samtools (experiments were run with v1.17)

Sequence-to-graph alignment tools for comparison

• PLAST (commit 451369eb23e84328ff9334398d185798d2dc5149)
• GraphAligner (v1.0.17b)

Additional software for evaluating alignment results

• WGSUniFrac (commit d5c698ba4699aba168fd29fc00024d655c36183f, included here)
• numpy (v1.26.0)
• pandas (v.2.1.1)

Data used for evaluations

• https://public.bmi.inf.ethz.ch/resources/mla/

Setting up the environment

1. To install most of the required software for the evaluation, set up a conda environment using the provided environment.yml file: conda env create -f environment.yml
2. Activate the environment: conda activate mla
3. Download the pre-compiled binaries of metagraph, PLAST, kmc, and parse_plast from here and place them in your working directory.
4. Download the genomes from here to a directory named references. A list of accessions is in accession_list
5. Download the random entropy source file seed2 (used to generate query sets)
6. Download the accession-ID augmented taxonomic tree from the augmented directory here

Constructing a simulated joint assembly graph and the query sets

1. Simulate reads for each genome:

   for a in references/*.fa; do ./make_sample.sh $a; done
   for a in illumina hifi clr ont; do ./make_subset.sh $a; done
   

2. Build the MetaGraph, PLAST, and GFA indexes by running ./make_graph.sh. In this script, you can set the number of threads in the variable $NTHREADS. We provide precomputed indexes for MetaGraph here and for PLAST here.
3. Generate query reads by running ./make_subset.sh

Run the alignments and classify the reads

for a in query_reads/*.fa; do
    ./map_query_sca.sh $a
    ./map_query_mla.sh $a
    ./map_query_plast.sh $a
    ./map_query_ga.sh $a
done

Notes

• To compile some of the software yourself from source
  • parse_plast.cpp: we have provided a Makefile.
  • PLAST: we have provided a template CMakeLists.txt file in plast_cmake. Please edit it to point to the include directory and static lib of Bifrost.
  • MetaGraph (with packaged kmc): follow the instructions here. Remember to checkout the commit listed above from the hm/aln_alt_label_change branch before compiling.
• Run metagraph align to view the help menu listing alignment parameters. To list a more advanced set of parameters, run metagraph align --advanced.