Search

As whole genome sequencing (WGS) uncovers variants associated with rare and common diseases, an immediate challenge is to minimize false-positive findings due to sequencing and variant calling errors. False positives can be reduced by combining results from orthogonal sequencing methods, but costly. Here, we present variant filtering approaches using logistic regression (LR) and ensemble genotyping to minimize false positives without sacrificing sensitivity. We evaluated the methods using paired WGS datasets of an extended family prepared using two sequencing platforms and a validated set of variants in NA12878. Using LR or ensemble genotyping based filtering, false-negative rates were significantly reduced by 1.1- to 17.8-fold at the same levels of false discovery rates (5.4% for heterozygous and 4.5% for homozygous single nucleotide variants (SNVs); 30.0% for heterozygous and 18.7% for homozygous insertions; 25.2% for heterozygous and 16.6% for homozygous deletions) compared to the filtering based on genotype quality scores. Moreover, ensemble genotyping excluded > 98% (105,080 of 107,167) of false positives while retaining > 95% (897 of 937) of true positives in de novo mutation (DNM) discovery in NA12878, and performed better than a consensus method using two sequencing platforms. Our proposed methods were effective in prioritizing phenotype-associated variants, and an ensemble genotyping would be essential to minimize false-positive DNM candidates., (© 2014 WILEY PERIODICALS, INC.)

Whole-genome sequencing (WGS) studies are uncovering disease-associated variants in both rare and nonrare diseases. Utilizing the next-generation sequencing for WGS requires a series of computational methods for alignment, variant detection, and annotation, and the accuracy and reproducibility of annotation results are essential for clinical implementation. However, annotating WGS with up to date genomic information is still challenging for biomedical researchers. Here, we present one of the fastest and highly scalable annotation, filtering, and analysis pipeline-gNOME-to prioritize phenotype-associated variants while minimizing false-positive findings. Intuitive graphical user interface of gNOME facilitates the selection of phenotype-associated variants, and the result summaries are provided at variant, gene, and genome levels. Moreover, the enrichment results of specific variants, genes, and gene sets between two groups or compared with population scale WGS datasets that is already integrated in the pipeline can help the interpretation. We found a small number of discordant results between annotation software tools in part due to different reporting strategies for the variants with complex impacts. Using two published whole-exome datasets of uveal melanoma and bladder cancer, we demonstrated gNOME's accuracy of variant annotation and the enrichment of loss-of-function variants in known cancer pathways. gNOME Web server and source codes are freely available to the academic community (http://gnome.tchlab.org)., (© 2014 WILEY PERIODICALS, INC.)