example.md

Example use case

This example is written for Themisto v3.0.0 or newer and mSWEEP v2.0.0 or newer.

Downloading the data

Download a toy dataset from Zenodo.

Indexing

Create a list containing paths to the input assemblies

ls -d $(pwd)"/assemblies/"*.fasta.gz > input_sequences.txt

Index the input data with Themisto (v3.0.0 or newer)

mkdir tmp
themisto build -k 31 -i input_sequences.txt -o themisto_index --temp-dir tmp -t 2 --mem-gigas 4

this will create the themisto_index.tcolors and themisto_index.tdbg files.

(optional) Read correction

Correct errors in the reads using [fastp](https://github.com/opengene/fastp]

fastp --in1 215_1.fastq.gz --in2 215_2.fastq.gz --out1 corr_1.fastq.gz --out2 corr_2.fastq.gz -c --thread 2

Pseudoalignment

Pseudoalign the reads with Themisto

themisto pseudoalign -q 215_1.fastq.gz -i themisto_index --temp-dir tmp -t 2 > 215_1.txt
themisto pseudoalign -q 215_2.fastq.gz -i themisto_index --temp-dir tmp -t 2 > 215_2.txt

(optional) pseudoalign and compress alignment

Pseudoalign the reads as above and compress the alignment file with alignment-writer

ntargets=$(wc -l clustering.txt | cut -f1 -d' ')
nreads=$((`gunzip -c 215_1.fastq.gz | wc -l` / 4 ))
themisto pseudoalign -q 215_1.fastq.gz -i themisto_index --temp-dir tmp -t 1 | alignment-writer -n $ntargets -r $nreads > 215_1.aln
themisto pseudoalign -q 215_1.fastq.gz -i themisto_index --temp-dir tmp -t 1 | alignment-writer -n $ntargets -r $nreads > 215_2.aln

this is particularly useful for very large alignments. mSWEEP v2.0.0 and newer can automatically detect the file format if the alignment files are compressed.

Abundance estimation

Estimate abundances with

mSWEEP --themisto-1 215_1.txt --themisto-2 215_2.txt -i clustering.txt -t 2

this will print the abundances after the estimation is finished. To write the abundances to 215_abundances.txt, run

mSWEEP --themisto-1 215_1.txt --themisto-2 215_2.txt -i clustering.txt -t 2 -o 215

Binning and abundance estimation

Bin the reads by adding the --bin-reads toggle

mSWEEP --themisto-1 215_1.txt --themisto-2 215_2.txt -i clustering.txt -t 2 --bin-reads

which will create the clust1.bin, clust2.bin, clust3.bin, and clust4.bin files. These files can be used as input to mGEMS to extract the reads from the input data to themisto pseudoalign.