LoFreq (i.e. LoFreq version 2) is a fast and sensitive variant-caller for inferring SNVs and indels from next-generation sequencing data. It makes full use of base-call qualities and other sources of errors inherent in sequencing (e.g. mapping or base/indel alignment uncertainty), which are usually ignored by other methods or only used for filtering.

LoFreq can run on almost any type of aligned sequencing data (e.g. Illumina, IonTorrent or Pacbio) since no machine- or sequencing-technology dependent thresholds are used. It automatically adapts to changes in coverage and sequencing quality and can therefore be applied to a variety of data-sets e.g. viral/quasispecies, bacterial, metagenomics or somatic data.

LoFreq is very sensitive; most notably, it is able to predict variants below the average base-call quality (i.e. sequencing error rate). Each variant call is assigned a p-value which allows for rigorous false positive control. Even though it uses no approximations or heuristics, it is very efficient due to several runtime optimizations and also provides a (pseudo-)parallel implementation. LoFreq* is generic and fast enough to be applied to high-coverage data and large genomes. On a single processor it takes a minute to analyze Dengue genome sequencing data with nearly 4000X coverage, roughly one hour to call SNVs on a 600X coverage E.coli genome and also roughly an hour to run on a 100X coverage human exome dataset.

References:

• http://www.ncbi.nlm.nih.gov/pubmed/23066108

• http://csb5.github.io/lofreq/commands/

• Module Name: lofreq (see the modules page for more information)

Allocate an interactive session and run the program. Sample session:

[user@biowulf]$ sinteractive
salloc.exe: Pending job allocation 46116226
salloc.exe: job 46116226 queued and waiting for resources
salloc.exe: job 46116226 has been allocated resources
salloc.exe: Granted job allocation 46116226
salloc.exe: Waiting for resource configuration
salloc.exe: Nodes cn3144 are ready for job

[user@cn3144 ~]$ module load lofreq
[user@cn3144 ~]$ lofreq call -f ref.fa -o vars.vcf aln.bam

[user@cn3144 ~]$ exit
salloc.exe: Relinquishing job allocation 46116226
[user@biowulf ~]$

Create a batch input file (e.g. batch.sh). For example:

#!/bin/bash
set -e
module load lofreq
lofreq call -f ref.fa -o vars.vcf aln.bam

Submit this job using the Slurm sbatch command.

sbatch [--mem=#] batch.sh

Create a swarmfile (e.g. job.swarm). For example:

cd dir1; lofreq call -f ref.fa -o vars.vcf aln.bam
cd dir2; lofreq call -f ref.fa -o vars.vcf aln.bam
cd dir3; lofreq call -f ref.fa -o vars.vcf aln.bam

Submit this job using the swarm command.

swarm -f job.swarm [-g #] --module lofreq