PLASS - Protein-Level ASSembler
Plass (Protein-Level ASSembler) is a software to assemble short read sequencing data on a protein level. The main purpose of Plass is the assembly of complex metagenomic datasets. It assembles 10 times more protein residues in soil metagenomes than Megahit. Plass is GPL-licensed open source software that is implemented in C++ and available for Linux and macOS. The software is designed to run on multiple cores. Plass was used to create a Soil Reference Catalog (SRC) and a Marine Eukaryotic Reference Catalog (MERC).
Soil Reference Catalog (SRC) and Marine Eukaryotic Reference Catalog (MERC)
SRC was created by assembling 640 soil metagenome samples. MERC was assembled from the the metatranscriptomics datasets created by the TARA ocean expedition. Both catalogues were redundancy reduced to 90% sequence identity at 90% coverage. Each catalog is a single FASTA file containing the sequences, the header identifiers contain the Sequence Read Archive (SRA) identifiers. The catalogues can be downloaded here. We provide a HH-suite3 database called "BFD" containing sequences from the Metaclust, SRC, MERC and Uniport at here.
Install Plass
Plass can be install via conda or as statically compiled Linux version. Plass requires a 64-bit Linux/MacOS system (check with uname -a | grep x86_64) with at least the SSE2 instruction set.
# install from bioconda
conda install -c conda-forge -c bioconda plass
# static build with AVX2 (fastest)
wget https://mmseqs.com/plass/plass-linux-avx2.tar.gz; tar xvfz plass-linux-avx2.tar.gz; export PATH=$(pwd)/plass/bin/:$PATH
# static build with SSE4.1
wget https://mmseqs.com/plass/plass-linux-sse41.tar.gz; tar xvfz plass-linux-sse41.tar.gz; export PATH=$(pwd)/plass/bin/:$PATH
# static build with SSE2 (slowest, for very old systems)
wget https://mmseqs.com/plass/plass-linux-sse2.tar.gz; tar xvfz plass-linux-sse2.tar.gz; export PATH=$(pwd)/plass/bin/:$PATH
How to assemble
Plass can assemble both paired-end reads (FASTQ) and single reads (FASTA or FASTQ):
# assemble paired-end reads
plass assemble examples/reads_1.fastq.gz examples/reads_2.fastq.gz assembly.fas tmp
# assemble single-end reads
plass assemble examples/reads_1.fastq.gz assembly.fas tmp
# assemble single-end reads using stdin
cat examples/reads_1.fastq.gz | plass assemble stdin assembly.fas tmp
Important parameters:
--min-seq-id Adjusts the overlap sequence identity threshold
--min-length minimum codon length for ORF prediction (default: 40)
-e E-value threshold for overlaps
--num-iterations Number of iterations of assembly
--filter-proteins Switches the neural network protein filter off/on
Modules:
plass assemble Assembles proteins (i:Nucleotides -> o:Proteins)
plass nuclassemble Assembles nucleotides *experimental* (i:Nucleotides -> o:Nucleotides)
Assemble using MPI
Plass can be distrubted over several homogeneous computers. However the TMP folder has to be shared between all nodes (e.g. NFS). The following command assembles several nodes:
RUNNER="mpirun -np 42" plass assemble examples/reads_1.fastq.gz examples/reads_2.fastq.gz assembly.fas tmp
Compile from source
Compiling PLASS from source has the advantage that it will be optimized to the specific system, which should improve its performance. To compile PLASS git, g++ (4.6 or higher) and cmake (3.0 or higher) are required. Afterwards, the PLASS binary will be located in the build/bin directory.
git clone https://github.com/soedinglab/plass.git
cd plass
git submodule update --init
mkdir build && cd build
cmake -DCMAKE_BUILD_TYPE=RELEASE -DCMAKE_INSTALL_PREFIX=. ..
make -j 4 && make install
export PATH="$(pwd)/bin/:$PATH"
gcc from Homebrew. The default macOS clang compiler does not support OpenMP and PLASS will not be able to run multithreaded. Use the following cmake call:
CXX="$(brew --prefix)/bin/g++-8" cmake -DCMAKE_BUILD_TYPE=RELEASE -DCMAKE_INSTALL_PREFIX=. ..
Dependencies
When compiling from source, PLASS requires zlib and bzip.
Use the docker image
We also provide a Docker image of Plass. You can mount the current directory containing the reads to be assembled and run plass with the following command:
docker pull soedinglab/plass
docker run -ti --rm -v "$(pwd):/app" -w /app plass assemble reads_1.fastq reads_2.fastq assembly.fas tmp
Hardware requirements
Plass needs roughly 1 byte of memory per residue to work efficiently. Plass will scale its memory consumption based on the available main memory of the machine. Plass needs a CPU with at least the SSE4.1 instruction set to run.
Known problems
- The assembly of Plass includes all ORFs having a start and end codon that includes even very short ORFs < 60 amino acids. Many of these short ORFs are spurious since our neural network cannot distingue them well. We would recommend to use other method to verify the coding potential of these. Assemblies above 100 amino acids are mostly genuine protein sequences.
- Plass in default searches for ORFs of 40 amino acids or longer. This limits the read length to > 120. To assemble this protein, you need to lower the
--min-lengththreshold. Be aware using short reads (< 100 length) might result in lower sensitivity.