ULLAR

ULLAR, named after ular, which means snakes in the Indonesian language, stands for an Ultrafast, scaLable, Accessible, and Reproducible pipeline for phylogenomics. Our goal with ULLAR is to develop a lightweight and scalable pipeline that requires a minimal learning curve. In addition to Linux and MacOS, the typical supported operating systems for bioinformatics, whenever possible, ULLAR will run natively on Windows.

Development Status

ULLAR is currently under development. We are working on the pipeline's core components. You should expect command changes in the future release. If you use ULLAR in publication, we recommend stating the exact version of the app. For manual compilation, we recommend to also state the commit hash number. For example, ULLAR v0.3.0 (commit: f18ac98).

Try ULLAR

You can try the pipeline by following the installation guide below. This guideline assume familiarity of using command line app and basic bioinformatics tools.

Installation

Currently, ULLAR installation requires Rust. Follow Rust installation guide here. After installing Rust, you can install ULLAR using cargo:

cargo install --git https://github.com/hhandika/ullar.git

Another option is to install ULLAR pre-compiled binary. You can download the latest release from the release page. Available binaries:

OS	Download
Linux	Intel/AMD 64-bit or Many Linux Intel/AMD 64-bit
Windows	Intel/AMD 64-bit
MacOS	Intel or M series

Install ULLAR like installing any single executable binary. For example, in Linux:

tar -xvf ullar-Linux-x86_64.tar.gz

Copy to your bin directory such as /usr/local/bin:

sudo cp ullar /usr/local/bin

or our home directory that is in the PATH if you don't have root access:

cp ullar ~/bin

SEGUL provide a detailed installation guide on installing Rust based software here

Features & Dependencies

Feature	Dependencies
Raw read cleaning	Fastp
De novo assembly	SPAdes
Reference mapping	LASTZ
Sequence alignment	MAFFT
ML phylogeny (in development)	IQ-TREE
MSC phylogeny (in development)	ASTER
Data cleaning	SEGUL
Summary statistics (in development)	SEGUL

NOTE: Summary statistics and other data cleaning feature is under development, but you can install SEGUL separately. Check out SEGUL documentation here

You can check if you have the dependencies installed by running the following commands:

ullar deps check

If you don't have the dependencies installed, you can install by following the instructions on the links provided above.

Check ULLAR installation:

ullar --version

Cleaning raw reads

A quick way to clean reads:

ullar clean init -d /raw_read_dir --autorun

By default ullar use descriptive name format to match the sample. It is equal to running ullar using this argument:

ullar clean init -d /raw_read_dir --sample-name descriptive

Example of descriptive names:

- sample1_Species1_R1.fastq.gz
- sample1_Species1_R2.fastq.gz
- genus1_species1_locality_R1.fastq.gz
- genus1_species1_locality_R2.fastq.gz
- genus1_species2_locality_R1.fastq.gz
- genus1_species2_locality_R2.fastq.gz

If your file naming is simple, you can use the --sample-name simple argument:

ullar clean init -d /raw_read_dir --sample-name simple

Example of simple names:

- sample1_R1.fastq.gz
- sample1_R2.fastq.gz

You can also supply your own regular expression to extract the sample name:

ullar clean init -d /raw_read_dir --re-sample='([a-zA-Z0-9]+)_R1.fastq.gz'

To run the cleaning process:

ullar clean run -c configs/read_cleaning.yaml

If you prefer to check the config file before running the cleaning process, you can init ullar without the --autorun argument:

ullar clean init -d /raw_read_dir

To run the cleaning process, you can use the --skip-config-check argument to skip the config check:

ullar clean run -c configs/read_cleaning.yaml

De Novo Assembly

ULLAR uses SPAdes for de novo assembly. To run the assembly:

ullar assemble init -d /cleaned_read_dir --autorun

If you prefer to check the config file before running the assembly process, you can init ullar without the --autorun argument:

ullar assemble init -d /cleaned_read_dir

To run the assembly process, you can use the --skip-config-check argument to skip the config check:

ullar assemble run -c configs/de_novo_assembly.yaml

Reference Mapping

ULLAR uses LASTZ for reference mapping. To run the mapping:

ullar map init -d /path/to/cleaned_read_dir --reference /path/to/reference.fasta --reference-type probes --autorun

If your reference is a locus, you can use the --reference-type loci argument:

ullar map init -d /path/to/cleaned_read_dir --reference /path/to/reference.fasta --reference-type loci --autorun 

For the probes type, ULLAR will pull an entire contig that matches the probe. The output will in lastz general format and sequence files in FASTA format.

For the loci type, ULLAR will only pull the part of the contig that matches the reference. The output will be in Multi Alignment Format (MAF) and FASTA format.

Sequence Alignment

ULLAR uses MAFFT for sequence alignment. To run the sequence alignment:

ullar align init -d /path/to/unaligned_sequences_dir --autorun

If you prefer to check the config file before running the alignment process, you can init ullar without the --autorun argument:

ullar align init -d /path/to/unaligned_sequences_dir

To run the alignment process, you can use the --skip-config-check argument to skip the config check:

ullar align run -c configs/sequence_alignment.yaml

Phylogenetic Analysis

ULLAR uses IQ-TREE for phylogenetic analysis. To run the phylogenetic analysis:

ullar tree init -d /path/to/aligned_sequences_dir --autorun

If you prefer to check the config file before running the phylogenetic analysis process, you can init ullar without the --autorun argument:

ullar tree init -d /path/to/aligned_sequences_dir

To run the phylogenetic analysis process, you can use the --skip-config-check argument to skip the config check:

ullar tree run -c configs/phylogenetic_analysis.yaml

To specify the tree inference method, you can use the --specify-analyses argument:

Options are:

• ml-species for maximum likelihood species tree inference
• ml-genes for maximum likelihood gene tree inference
• gscf for gene site concordance factor
• msc for multi-species coalescent tree inference

For example:

ullar tree init -d /path/to/aligned_sequences_dir --specify-analyses ml-species ml-genes --autorun

You can also specify the multi-species coalescent tree inference method using --specify-msc-methods argument:

Options are:

• astral for ASTRAL 4 methods.
• astral-pro for ASTRAL Pro methods.
• wastral for Weighted ASTRAL methods.

MSC methods requires --specify-analyses ml-genes msc argument.

For example:

ullar tree init -d /path/to/aligned_sequences_dir --specify-analyses ml-genes msc --specify-msc-methods astral-pro --autorun