Following are the features supported by the latest verison of Varant:

Features	Descriptions
Variant type that the tool annotates	SNPs, Indels & MNPs
Input file format	VCF
Output file format	VCF
VCF Parser	Provides parser for Varant annotated VCF

Following are the 22 annotations provided by Varant:

1. dbSNP, 1000Genome Minor Allele Frequency (MAF) & ESP (MAF)
2. Clinically significant variants from ClinVar DB
3. GWAS Phenotype
4. Genomic region - Intergenic, Intronic, Exonic & UTR
5. Downstream and upstream gene for intergenic variants
6. Splice Site (Donor/Acceptor)
7. Mutation Type - NonSyn, Syn, StartGain, StartLoss, StopGain, StopLoss, SynStop
8. Codon Usage in Human
9. Exonic splice enhancer / silencer site
10. Flag variants that spans boundary region like Intron-Exon or UTR-CDS
11. Distance of intronic variants from splice sites
12. UTR Functional Motifs
13. miRNA Binding Site
14. Polyphen2, SIFT & CADD prediction
15. Gene-Disease association - OMIM, NCBI-GAD
16. Position Conservation - Gerp++ Score
17. Interpro Domain
18. TFBS
19. eQTL
20. Low complexity region
21. Pseudo autosomal region
22. Capture region

Varant annotations are written to the INFO field of the VCF file in compliance to the VCF file format and can be easily parsed by any VCF parser.

Following are the details about the format -

dbSNP, 1000Genome or ESP variants
Clinically significant variants from ClinVar DB
GWAS variants
Intergenic variants
Genic variants
Conserved Regions
CADD prediction
Interpro Domain
TFBS and eQLT
Low complexity regions
Pseudo Autosomal regions
Capture regions

dbSNP, 1000Genome or ESP variants

Annotations in INFO field of VCF file	Description
DB138	Variant present in dbSNP138
dbSNPBuildID	dbSNP build ID in which the variant was first reported
KGDB	Variant present in 1000Genome
KGAF	The minor allele frequency (fraction) reported in 1000Genome. Eg KGAF=0.002
ESPDB	Variant present in Exome Sequencing Project
ESPAF	The minor allele frequency (fraction) reported in Exome sequencing project. Eg ESPAF=0.01

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 24180962 rs13551 T C 100.0 PASS DB138;ESPAF=0.268184;ESPDB;KGAF=0.25;KGDB;dbSNPBuildID=52

Clinically significant variants from ClinVar DB

Annotations in INFO field of VCF file	Description
CLNDSDB	Variant disease database name
CLNACC	Variant Accession and Versions
CLNDBN	Variant disease name
CLNSRC	Variant Clinical Chanels
CLNSIG	Variant Clinical Significance, 0 - unknown, 1 - untested, 2 - non-pathogenic, 3 - probable-non-pathogenic, 4 - probable-pathogenic, 5 - pathogenic, 6 - drug-response, 7 - histocompatibility, 255 - other
CLNORIGIN	Allele Origin. One or more of the following values may be added: 0 - unknown; 1 - germline; 2 - somatic; 4 - inherited; 8 - paternal; 16 - maternal; 32 - de-novo; 64 - biparental; 128 - uniparental; 256 - not-tested; 512 - tested-inconclusive; 1073741824 - other
CLN	SNP is Clinical(LSDB,OMIM,TPA,Diagnostic)
CLNDSDBID	Variant disease database ID
CLNHGVS	Variant names from HGVS. The order of these variants corresponds to the order of the info in the other clinical INFO tags.
CLNSRCID	Variant Clinical Channel IDs

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 1959699 rs41307846 G A 100.0 PASS CLNACC=RCV000017599.1|RCV000017600.1|RCV000022558.1;CLNDBN=Generalized_epilepsy_with_febrile_seizures_plus_type_5|Epilepsy\x2c_juvenile_myoclonic_7|Epilepsy\x2c_idiopathic_generalized_10;CLNDSDB=MedGen|MedGen|MedGen:OMIM;CLNDSDBID=C3150401|CN043549|C2751603:613060;CLNHGVS=NC_000001.10:g.1959699G>A;CLNORIGIN=1;CLNSIG=255|255|255;CLNSRC=OMIM_Allelic_Variant;CLNSRCID=137163.0002

GWAS variants

Annotations in INFO field of VCF file	Description
GWASPhenotype	NHGRI-GWAS phenotypes associated with the variant

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 15 74219582 rs3825942 G A . . GWASPhenotype=Glaucoma_exfoliation

Intergenic variants

For the variants in intergenic region, the downstream and upstream genes along with the distance from the variant to the gene’s TSS/TES are reported in the following format –

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 1264463 rs307353 A G 100.0 PASS VARANT_INTERGENIC=GLTPD1(dist=187):TAS1R3(dist=2263)

Genic variants

Variant in genic region is annotated in following the format -

If there are more than one transcript for a gene, then the annotations on them are appended after the annotations for the first transcript by ':'. The last three annotations appended by ':' to the transcript related annotations are always OMIM phenotype, OMIM phenotype Ids and GAD phenotypes associated with the gene.

Following is the description of the fields in VARANT_GENIC :

Fields	Description
Gene	Gene Name
Transcript_ID	Transcript Accession number
Region	Genomic region where the variant is present. Following are the possible values for this field -
	CodingExonic - Variant present in CDS
	CodingIntronic - Variant present in intron of coding transcript
	NonCodingExonic - Variant present in exon of non-coding transcript
	NonCodingIntronic - Variant present in intron of non-coding transcript
	UTR5 - Variant present in 5'UTR of transcript
	UTR3 - Variant present in 3'UTR of transcript
	Intergenic_UTR5_boundary - Variant spanning Intergenic and 5'UTR region
	CodingExonic_CodingIntronic_boundary - Variant spanning CDS and intronic region of coding transcript
	UTR5_CodingExonic_boundary - Variant spanning 5'UTR and CDS of coding transcript
	CodingExonic_UTR3_boundary - Variant spanning CDS and 3'UTR of coding transcript
	UTR3_Intergenic_boundary - Variant spanning 3'UTR and Intergenic region of coding transcript
	NonCodingExonic_NonCodingIntronic_boundary - Variant spanning exon and intron of non-coding transcript
Exon_Number	Exon which hosts the variant. If the variant spans more than one exon then the exon numbers are reported as '__' delimited.
AltID	Alternate allele id number (1,2,3 etc) to which the annotations corresponds
cDNAPos	cDNA position of the variant
SpliceSite	Splicing site or splice regulation site annotations. Following are the possible values for this field -
	ESE - Variant present in predicted Exonic Splice Enhancer Site
	ESS - Variant present in predicted Exonic Splice Silencer Site
	SpliceDonor - Variant present at 5' splice site
	SpliceAcceptor - Variant present at 3' splice site
	ExonX_SpliceDonor_dist1__ExonY_SpliceAcceptor_dist2 - This annotation is reported for the intronic variants not at splice sites. The ExonX and ExonY are the flanking exons with X and Y as exon numbers, dist1 is the distance of the variant from SpliceDonor site and dist2 is the distance of the variant from SpliceAcceptor site. E.g. Exon7_SpliceDonor_5__Exon8_SpliceAcceptor_750 indicating the variant is 5 bp away from SpliceDonor and 750bp away from SpliceAcceptor and the variant is between Exon7 and Exon8
UTRSignal	Functional Motifs in the untranslated region of transcript
Mutation	Mutation type caused by variant. Following are the possible values for this field -
	NonSyn, Syn, StartGain, StartLoss, StopGain, StopLoss, FrameShiftInsert, FrameShiftDelete, NonFrameShiftInsert, NonFrameShiftDelete, NoCDSChange
Codon_Change	Reports wild type codon and mutant type codon that hosts the variant
Amino_Acid_Change	Change in the amino acid due to mutation. Reports the wild type and mutant type amino acids and protein position
Ref_Protein_Length	The original protein length
Codon_Usage	Change in codon usage. Following are the possible values for this field -
	CodonUsageDown - The mutant type codon usage is lower than the wild type codon usage
	CodonUsageUp - The mutant type codon usage is higher than the wild type codon usage
SIFT(pred_score)	The SIFT prediction and score for the NonSyn mutations. Following are the possible values for this field -
	T_Score - 'T' stands for Tolerated effect followed by SIFT score. E.g. T_0.28
	D_Score - 'D' stands for Damaging effect followed by SIFT score. E.g. D_0.02
Polyphen2(pred_score)	The Polyphen2 prediction and score for the NonSyn mutations. Following are the possible values for this field -
	PP2B_Score - 'PP2B' stands for Benign effect followed by PolyPhen2 score. E.g. PP2B_0.018
	PP2PD_Score - 'PP2PD' stands for Possibly Damaging effect followed by PolyPhen2 score. E.g. PP2PD_0.789
	PP2D_Score - 'PP2D' stands for Probably Damaging effect followed by PolyPhen2 score. E.g. PP2D_0.991
Warning	Annotation Warnings and following are the possible values for this field -
	NOT_ACTUAL_STOP_CODON__TRANSCRIPT_WITH_MULTIPLE_STOP_CODON - This warning is reported when there is a StopLoss mutation and the stop codon is not present at the end of CDS.
	CDS_NOT_MULTIPLE_OF_3 - This warning is reported when the transcript is incomplete.
OMIM_Disease	Gene associated OMIM Phenotypes. Mutiple values are '__' delimited.
OMIM_Ids	Gene associated OMIM Phenotype IDs. Mutiple values are '__' delimited.
GAD_Disease	Gene associated Phenotypes from NCBI-GAD database. Mutiple values are '__' delimited.

Example:
Variant in CDS of a gene and causing NonSyn mutation and predicted as damaging. The gene is also associated with a OMIM phenotype.
#CHROM POS ID REF ALT QUAL FILTER INFO 1 53712727 rs5174 C T 100.0 PASS VARANT_GENIC=LRP8(NM_004631|CodingExonic|1|1|3092|||NonSyn|CGA/CAA|R952Q|963|CodonUsageUp|D_0.05|PP2D_0.999|:NM_017522|CodingExonic|1|1|2303|||NonSyn|CGA/CAA|R689Q|700|CodonUsageUp|D_0.05|PP2D_1.0|:NM_033300|CodingExonic|1|1|2582|||NonSyn|CGA/CAA|R782Q|793|CodonUsageUp|D_0.05|PP2D_0.999|:NM_001018054|CodingExonic|1|1|2915|||NonSyn|CGA/CAA|R893Q|904|CodonUsageUp|D_0.05|PP2PD_0.832|:MYOCARDIAL_INFARCTION_SUSCEPTIBILITY_TO:608446:birth_weight_fetal_growth)
Variant that spans boundary region.
#CHROM POS ID REF ALT QUAL FILTER INFO 19 36317406 rs386833941 GAGAGGGCTCTTACACC G . . VARANT_GENIC=NPHS1(NM_004646|CodingExonic_UTR3_boundary|1|1|3876_3877_3878_3879_3880_3881_3882_3883_3884_3885_3886_3887_3888_3889_3890_3891|||StopLoss|||||||:NEPHROTIC_SYNDROME_TYPE_1:256300:Ig_A_nephropathy__nephrotic_syndrome__nephropathy)
Variant not at splice site but very close to 5' splice site.
#CHROM POS ID REF ALT QUAL FILTER INFO 1 201335960 rs397516449 A G . . VARANT_GENIC=TNNT2(NM_001276347|CodingIntronic||1||Exon9_SpliceDonor_4__Exon10_SpliceAcceptor_1160|||||||||:CARDIOMYOPATHY_FAMILIAL_HYPERTROPHIC_2__CARDIOMYOPATHY_FAMILIAL_RESTRICTIVE_3__CARDIOMYOPATHY_DILATED_1D:115195__612422__601494:cardiomyopathy__left_ventricular_hypertrophy)
Variant on overlapping genes.
#CHROM POS ID REF ALT QUAL FILTER INFO 3 181430369 rs104893805 G C . . VARANT_GENIC=SOX2(NM_003106|CodingExonic|1|1|658|ESS||NonSyn|CGC/CCC|R74P|317||D_0.0|PP2PD_0.916|:MICROPHTHALMIA_SYNDROMIC_3:206900:),SOX2-OT(NR_004053|NonCodingIntronic||1||Exon3_SpliceDonor_12696__Exon4_SpliceAcceptor_2602|||||||||:::)

Conserved Regions

Annotations in INFO field of VCF file	Description
GerpConserve	Flags the variant to be present in a conserved region
GerpRSScore	Gerp Rejected Substitutions Score. Rejected substitutions are a natural measure of constraint that reflects the strength of past purifying selection on the element.
GerpPValue	Gerp P-Value

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 891344 rs267598748 G A . . GerpConserve;GerpPValue=3.65828e-36;GerpRSScore=182.5

CADD prediction

Annotations in INFO field of VCF file	Description
CADD_raw	CADD raw score for funtional prediction of a SNP. The larger the score the more likely the SNP has damaging effect. Scores are reported in the order in which ALT alleles are reported.
CADD_phred	CADD phred-like score. This is phred-like rank score based on whole genome CADD raw scores. The larger the score the more likely the SNP has damaging effect. Scores are reported in the order in which ALT alleles are reported

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 976598 . C T 7.83879 QUALFILTER CADD_phred=19.7;CADD_raw=3.876027

Interpro Domain

Annotations in INFO field of VCF file	Description
Interpro_domains	Variant position is part of a domain or conserved site

Example: #CHROM POS ID REF ALT QUAL FILTER INFO 1 976700 . A C 0.00932919 QUALFILTER Interpro_domains=Proteaseinhibitor_Kazal-type,ProteinaseinhibitorI1_Kazal

TFBS and eQLT

Regulome DB is used for this annotation.

Annotations in INFO field of VCF file	Description
RegulomeScore	Regulome Score as described here . Following are the possible values for this field -
	1a or 1b or 1c or 1d or 1e or 1f - Represents eQTL
	2a or 2b or 2c or 3a or 3b or 4 - Represents TFBS

Example:
Variant upstream of a gene and at TFBS
#CHROM POS ID REF ALT QUAL FILTER INFO 1 155113074 rs12745576 A C . . RegulomeScore=4
Variant causing Syn mutation but is a eQTL
#CHROM POS ID REF ALT QUAL FILTER INFO 7 100225031 rs2075674 G A . . RegulomeScore=1f

Low complexity regions

The LCR data used was distributed as supplement material for the paper - Towards Better Understanding of Artifacts in Variant Calling from High-Coverage Samples by Heng Li.
The file used for annotation was downloaded from - LCR-hs37d5.bed.gz

Annotations in INFO field of VCF file	Description
LCR	Variant position is part of low-complexity region

Example: An insertion variant in low complexity region
#CHROM POS ID REF ALT QUAL FILTER INFO 1 32403690 . T TGGC 0.0189515 QUALFILTER LCR

Pseudo Autosomal regions

The genes in PAR regions were compiled from the following two sources -
1. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2435358/
2. http://en.wikipedia.org/wiki/Pseudoautosomal_region

Annotations in INFO field of VCF file	Description
PAR	Variant is in Pseudoautosomal Region

Example: #CHROM POS ID REF ALT QUAL FILTER INFO X 299591 . A G 47091.7 TOOFREQUENTFILTER PAR

Capture regions

The current version is set up to annotate using Nimblegen capture bed file but this feature can be extended to use other capture arrays from Illumina or Agilent.

Annotations in INFO field of VCF file	Description
CaptureCore	Variant is present in Capture bed file
Capture5p	Number of bases that the variant position is 5p upstream of capture region
Capture3p	Number of bases that the variant position is 3p downstream of capture region

Example:
Variant is part of capture region #CHROM POS ID REF ALT QUAL FILTER INFO 1 69511 rs75062661 A G 28975.4 TOOFREQUENTFILTER CaptureCore
Variant position is 20 bases upstream of capture region #CHROM POS ID REF ALT QUAL FILTER INFO 1 1407165 rs140699979 C T 76.0097 PASS Capture5p=20
Variant position is 28 bases downstream of capture region #CHROM POS ID REF ALT QUAL FILTER INFO 1 15724003 rs2253371 A G 11310.3 TOOFREQUENTFILTER Capture3p=28