title: The struggle to find reliable results in exome sequencing data: filtering out Mendelian errors.
abstract: Next Generation Sequencing studies generate a large quantity of genetic data in a relatively cost and time efficient manner and provide an unprecedented opportunity to identify candidate causative variants that lead to disease phenotypes. A challenge to these studies is the generation of sequencing artifacts by current technologies. To identify and characterize the properties that distinguish false positive variants from true variants, we sequenced a child and both parents (one trio) using DNA isolated from three sources (blood, buccal cells, and saliva). The trio strategy allowed us to identify variants in the proband that could not have been inherited from the parents (Mendelian errors) and would most likely indicate sequencing artifacts. Quality control measurements were examined and three measurements were found to identify the greatest number of Mendelian errors. These included read depth, genotype quality score, and alternate allele ratio. Filtering the variants on these measurements removed ~95% of the Mendelian errors while retaining 80% of the called variants. These filters were applied independently. After filtering, the concordance between identical samples isolated from different sources was 99.99% as compared to 87% before filtering. This high concordance suggests that different sources of DNA can be used in trio studies without affecting the ability to identify causative polymorphisms. To facilitate analysis of next generation sequencing data, we developed the Cincinnati Analytical Suite for Sequencing Informatics (CASSI) to store sequencing files, metadata (eg. relatedness information), file versioning, data filtering, variant annotation, and identify candidate causative polymorphisms that follow either de novo, rare recessive homozygous or compound heterozygous inheritance models. We conclude the data cleaning process improves the signal to noise ratio in terms of variants and facilitates the identification of candidate disease causative polymorphisms.
authors.affiliation: Division of Rheumatology, Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA ; Medical Scientist Training Program, University of Cincinnati College of Medicine, Cincinnati OH, USA.
authors.forename: Zubin H
authors.initials: ZH
authors.lastname: Patel
authors.affiliation: Division of Rheumatology, Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA ; Department of Veterans Affairs, Veterans Affairs Medical Center - Cincinnati, Cincinnati OH, USA.
authors.forename: Leah C
authors.initials: LC
authors.lastname: Kottyan
authors.affiliation: Division of Rheumatology, Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA ; Department of Veterans Affairs, Veterans Affairs Medical Center - Cincinnati, Cincinnati OH, USA.
authors.forename: Sara
authors.initials: S
authors.lastname: Lazaro
authors.affiliation: Genomic Medicine Institute, Geisinger Health System, Danville PA, USA.
authors.forename: Marc S
authors.initials: MS
authors.lastname: Williams
authors.affiliation: Genomic Medicine Institute, Geisinger Health System, Danville PA, USA.
authors.forename: David H
authors.initials: DH
authors.lastname: Ledbetter
authors.affiliation: Genomic Medicine Institute, Geisinger Health System, Danville PA, USA.
authors.forename: Hbgerard
authors.initials: H
authors.lastname: Tromp
authors.affiliation: Division of Biomedical Informatics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Rupert
authors.affiliation: Division of Biomedical Informatics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Mojtaba
authors.initials: M
authors.lastname: Kohram
authors.affiliation: Division of Biomedical Informatics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Wagner
authors.affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Ammar
authors.initials: A
authors.lastname: Husami
authors.affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Yaping
authors.initials: Y
authors.lastname: Qian
authors.affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: C Alexander
authors.initials: CA
authors.lastname: Valencia
authors.affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Kejian
authors.initials: K
authors.lastname: Zhang
authors.affiliation: Division of Infectious Disease, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA.
authors.forename: Margaret K
authors.initials: MK
authors.lastname: Hostetter
authors.affiliation: Division of Rheumatology, Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA ; Department of Veterans Affairs, Veterans Affairs Medical Center - Cincinnati, Cincinnati OH, USA.
authors.forename: John B
authors.initials: JB
authors.lastname: Harley
authors.affiliation: Division of Rheumatology, Center for Autoimmune Genomics and Etiology, Cincinnati Children's Hospital Medical Center, Cincinnati OH, USA ; Department of Veterans Affairs, Veterans Affairs Medical Center - Cincinnati, Cincinnati OH, USA.
authors.forename: Kenneth M
authors.initials: KM
authors.lastname: Kaufman
keywords: CASSI; Mendel errors; Mendelian errors; disease causative polymorphisms; next-generation sequencing; variant filtering; whole exome sequencing