Assembly Metrics

Assess a genome assembly using various metrics (e.g. N50, total length, BUSCO)

Scripts found within the directory 01_scripts/00_utility_scripts originate from various sources, and these are attributed within each of the scripts themselves. Many are from Eric Normandeau's scripts repo: https://github.com/enormandeau/Scripts

Warning: this pipeline is provided "as is", without any warranty of any kind, express or implied.

Run scripts from the main directory of the repo.

Requirements:

Unix environment
biopython
BUSCO
bwa
samtools
python3

0. Prelim Options

You can confirm the smallest contig size using
./01_scripts/00_utility_scripts/fasta_min_len.py my_minl500.fasta

1. Simple Calculations on fasta

01_scripts/01_fasta_summarize.sh <assembly.fasta> <minimum_length>

Where minimum length is a number that is the smallest length that you would like your assembly to contain.

Provides for the full assembly and the limited length assembly:

• number of records
• total length
• n50

2. Markers against genome

First use bwa to index your assembly.fasta bwa index <assembly.fasta>

01_scripts/02_fasta_map.sh <query.fasta> <assembly.fasta>

Then use a utility script to get statistics from the .bam file
01_scripts/00_utility_scripts/bam_assess.sh 02_assemblies/<output.bam>

3. BUSCO evaluation

Currently only set up via a job submission script in batches, but can be adapted to single jobs from the script shown below. If it is of interest to adapt this to non-job submission, please let the author(s) know and it may be possible to include.

Using the job submission file, BUSCO can be run on multiple assemblies all found within 02_assemblies/*.fasta
To set up on your own directory, customize the following in the SLURM job file:

• Set path to BUSCO python script
• Set path to BUSCO lineage folder
• Set directories within the batch submission top part of the job script

After customizing to your directory, submit the job script
sbatch 01_scripts/jobs/job_BUSCO_multiple.sh

This will result in multiple BUSCO directories for each fasta file found in your 02_assemblies folder

Evaluate a metatranscriptome

1. Align fastq against metatranscriptome

To assess the quality of a metatranscriptome, one approach is to take several individual fastq files and see how well they map against the assembly. First index your metatranscriptome using bowtie2.

Then run the following command, including the bt2_index_base as <your_assembly>:
./01_scripts/03_aln_fa_to_ref_txome.sh 02_assemblies/your_assembly

The output results will be accessible in 04_mapping_results, both in .bam format saved under the name of the reference, as well as in log format.