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Genomic variant annotation and prioritization with ANNOVAR and wANNOVAR


Recent developments in sequencing techniques have enabled rapid and high-throughput generation of sequence data, democratizing the ability to compile information on large amounts of genetic variations in individual laboratories. However, there is a growing gap between the generation of raw sequencing data and the extraction of meaningful biological information. Here, we describe a protocol to use the ANNOVAR (ANNOtate VARiation) software to facilitate fast and easy variant annotations, including gene-based, region-based and filter-based annotations on a variant call format (VCF) file generated from human genomes. We further describe a protocol for gene-based annotation of a newly sequenced nonhuman species. Finally, we describe how to use a user-friendly and easily accessible web server called wANNOVAR to prioritize candidate genes for a Mendelian disease. The variant annotation protocols take 5–30 min of computer time, depending on the size of the variant file, and 5–10 min of hands-on time. In summary, through the command-line tool and the web server, these protocols provide a convenient means to analyze genetic variants generated in humans and other species.

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Figure 1: The three different types of annotations supported by ANNOVAR are gene-based, region-based and filter-based annotations.
Figure 2: Screenshot of wANNOVAR, including the general steps to upload and prioritize variants.
Figure 3: Screenshot of the wANNOVAR result page.


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Development of the ANNOVAR/wANNOVAR tool is supported by US National Institutes of Health (NIH) grant R01 HG006465. We thank X. Chang for the initial development of the wANNOVAR server. We thank all ANNOVAR and wANNOVAR users for their helpful suggestions, comments and bug reports to improve the software tools and web servers.

Author information




H.Y. and K.W. drafted and revised this manuscript.

Corresponding author

Correspondence to Kai Wang.

Ethics declarations

Competing interests

K.W. is a shareholder and board member of Tute Genomics.

Integrated supplementary information

Supplementary Figure 1 The expected results for discovering candidate genes of the ‘hemolytic anemia’ example in the Phenolyzer website.

Each ball represents one of the top 50 ranked genes. The larger the ball, the higher the ranking. The blue balls represent disease genes reported before and the yellow ones represent predicted disease genes. For detailed explanations on each color and shape, and on how the algorithm works to find disease genes, please visit

Supplementary Figure 2 Explanation of each column in the wANNOVAR web view.

This is a sample output with default parameters. The first 5 columns represent the original information on the input variants. The following 5 columns give gene-based annotations on each variant. The following columns give region-based and filter-based annotations on each variant.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1 and 2, Supplementary Tables 1–3 (PDF 841 kb)

Supplementary Data

ANNOVAR and VEP comparison results (XLSX 453 kb)

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Yang, H., Wang, K. Genomic variant annotation and prioritization with ANNOVAR and wANNOVAR. Nat Protoc 10, 1556–1566 (2015).

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