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snpEff is a variant annotation and effect prediction tool. It annotates and predicts the effects of variants on genes (such as amino acid changes).

Typical usage :

• Input: The inputs are predicted variants (SNPs, insertions, deletions and MNPs). The input file is usually obtained as a result of a sequencing experiment, and it is usually in variant call format (VCF).
• Output: SnpEff analyzes the input variants. It annotates the variants and calculates the effects they produce on known genes (e.g. amino acid changes). A list of effects and annotations that SnpEff can calculate can be found here.

References:

• A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3.", Cingolani P, Platts A, Wang le L, Coon M, Nguyen T, Wang L, Land SJ, Lu X, Ruden DM. Fly (Austin). 2012 Apr-Jun;6(2):80-92. PMID: 22728672

• snpEff Homepage
• snpSift Homepage
• snpEff Usage examples

• Module Name: snpEff (see the modules page for more information)
• Single-threaded, java-dependent
• environment variables set
  • $SNPEFF_HOME -- path to the installation directory
  • $SNPEFF_JAR -- the snpEff.jar file
  • $SNPEFF_JARPATH -- same as SNPEFF_HOME
  • $SNPSIFT_JAR -- the SnpSift.jar file
• Example files in $SNPEFF_HOME/../protocols
• Reference data in /fdb/dbNSFP2/

snpEff is a java application. See https://hpc.nih.gov/development/java.html for information about running java applications.

To see the help menu, type

java -jar $SNPEFF_JAR

at the prompt.

By default, snpEff uses 1gb of memory. For large VCF input files, this may not be enough. To allocate 20gb of memory, use:

java -Xmx20g -jar $SNPEFF_JAR

In more detail,

java -Xmx20g -jar $SNPEFF_JAR -v [ database ] [ vcf file ] 

To see what databases are available, type:

ls $SNPEFF_HOME/data

snpSift

SnpSift is a collection of tools to manipulate VCF (variant call format) files. Here's what you can do:

• Filter: You can filter using arbitrary expressions, for instance "(QUAL > 30) | (exists INDEL) | ( countHet() > 2 )". The actual expressions can be quite complex, so it allows for a lot of flexibility.
• Annotate: You can add 'ID' from another database (e.g. variants from dbSnp)
• CaseControl: You can compare how many variants are in 'case' and in 'control' groups. Also calculates p-values (Fisher exact test).
• Intervals: Filter variants that intersect with intervals.
• Intervals (intidx): Filter variants that intersect with intervals. Index the VCF file using memory mapped I/O to speed up the search. This is intended for huge VCF files and a small number of intervals to retrieve.
• Join: Join by generic genomic regions (intersecting or closest).
• RmRefGen: Remove reference genotype (i.e. replace '0/0' genotypes by '.')
• TsTv: Calculate transiton to transversion ratio.
• Extract fields: Extract fields from a VCF file to a TXT (tab separated) format.
• Variant type: Adds SNP/MNP/INS/DEL to info field. It also adds "HOM/HET" if there is only one sample.
• GWAS Catalog: Annotate using GWAS Catalog.
• dbNSFP: Annotate using dbNSFP: The dbNSFP is an integrated database of functional predictions from multiple algorithms (SIFT, Polyphen2, LRT and MutationTaster, PhyloP and GERP++, etc.)

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 snpEff
[user@cn3144 ~]$ ln -s $SNPEFF_HOME/../protocols .
[user@cn3144 ~]$ java -Xmx12g -jar $SNPEFF_JAR -v -lof -motif -hgvs -nextProt GRCh37.71 protocols/ex1.vcf > ex1.eff.vcf
[user@cn3144 ~]$ cat ex.eff.vcf | java -jar $SNPSIFT_JAR filter "(Cases[0] = 3) & (Controls[0] = 0) & ((EFF[*].IMPACT = 'HIGH') | (EFF[*].IMPACT = 'MODERATE'))"  > ex1.filtered.vcf
[user@cn3144 ~]$ java -jar $SNPSIFT_JAR dbnsfp -v -db /fdb/dbNSFP2/dbNSFP2.9.txt.gz ex1.eff.vcf > file.annotated.vcf

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

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

#!/bin/bash
# -- this file is snpEff.sh --

module load snpEff
ln -s $SNPEFF_HOME/example/file.vcf .
java -Xmx${SLURM_MEM_PER_NODE}m -jar $SNPEFF_JAR -v hg19 file.vcf > file.eff.vcf
cat file.eff.vcf | java -jar $SNPSIFT_JAR filter "( EFF[*].IMPACT = 'HIGH' )" > file.filtered.vcf
java -jar $SNPSIFT_JAR dbnsfp -v -db /fdb/dbNSFP2/dbNSFP3.2a.txt.gz file.eff.vcf > file.annotated.vcf

Submit this job using the Slurm sbatch command.

sbatch [--cpus-per-task=#] [--mem=#] snpEff.sh

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

java -Xmx${SLURM_MEM_PER_NODE}m -jar $SNPEFF_JAR -v hg19 file1.vcf > file1.eff.vcf
java -Xmx${SLURM_MEM_PER_NODE}m -jar $SNPEFF_JAR -v hg19 file2.vcf > file2.eff.vcf
java -Xmx${SLURM_MEM_PER_NODE}m -jar $SNPEFF_JAR -v hg19 file3.vcf > file3.eff.vcf
java -Xmx${SLURM_MEM_PER_NODE}m -jar $SNPEFF_JAR -v hg19 file4.vcf > file4.eff.vcf

Submit this job using the swarm command.

swarm -f snpEff.swarm [-g #] --module snpEff

where

-g #	Number of Gigabytes of memory required for each process (1 line in the swarm command file)
--module snpEff	Loads the snpEff module for each subjob in the swarm