BACKGROUND: Whole exome sequencing (WES) and whole genome sequencing (WGS) have become standard methods in human clinical diagnostics as well as in population genomics (POPGEN). Blood-derived genomic DNA (gDNA) is routinely used in the clinical environment. Conversely, many POPGEN studies and commercial tests benefit from easy saliva sampling. Here, we evaluated the quality of variant call sets and the level of genotype concordance of single nucleotide variants (SNVs) and small insertions and deletions (indels) for WES and WGS using paired blood- and saliva-derived gDNA isolates employing genomic reference-based validated protocols. METHODS: The genomic reference standard Coriell NA12878 was repeatedly analyzed using optimized WES and WGS protocols, and data calls were compared with the truth dataset published by the Genome in a Bottle Consortium. gDNA was extracted from the paired blood and saliva samples of 10 participants and processed using the same protocols. A comparison of paired blood-saliva call sets was performed in the context of WGS and WES genomic reference-based technical validation results. RESULTS: The quality pattern of called variants obtained from genomic-reference-based technical replicates correlates with data calls of paired blood-saliva-derived samples in all levels of tested examinations despite a higher rate of non-human contamination found in the saliva samples. The F1 score of 10 blood-to-saliva-derived comparisons ranged between 0.8030-0.9998 for SNVs and between 0.8883-0.9991 for small-indels in the case of the WGS protocol, and between 0.8643-0.999 for SNVs and between 0.7781-1.000 for small-indels in the case of the WES protocol. CONCLUSION: Saliva may be considered an equivalent material to blood for genetic analysis for both WGS and WES under strict protocol conditions. The accuracy of sequencing metrics and variant-detection accuracy is not affected by choosing saliva as the gDNA source instead of blood but much more significantly by the genomic context, variant types, and the sequencing technology used.

Comenius University Science Park Comenius University Ilkovičova 8 Karlova Ves Bratislava 841 04 Slovakia

Department of Botany and Zoology Faculty of Science Masaryk University Kotlářská 2 Brno 611 37 Czech Republic

Department of Molecular Biology Faculty of Natural Sciences Comenius University Ilkovičova 3278 6 Karlova Ves Bratislava 841 04 Slovakia

Faculty of Science Charles University Albertov 6 Prague 128 00 Czech Republic

Geneton s r o Ilkovičova 8 Bratislava 841 04 Slovakia

Institute of Applied Biotechnologies a s Služeb 4 Prague 108 00 Czech Republic

Institute of Biology and Medical Genetics 1st Faculty of Medicine Charles University and General University Hospital Prague Albertov 4 Prague 128 00 Czech Republic

Institute of Clinical and Translational Research Biomedical Research Center of the Slovak Academy of Sciences Dúbravská Cesta 9 Bratislava 845 05 Slovakia

Laboratory of Genomics and Bioinformatics Institute of Molecular Genetics of the Czech Academy of Sciences Vídeňská 1083 Prague 142 20 Czech Republic

Laboratory of Transcriptional Regulation Institute of Molecular Genetics of the Czech Academy of Sciences Vídeňská 1083 Prague 142 20 Czech Republic

Slovak Centre for Scientific and Technical Information Staré Mesto Lamačská Cesta 8A Bratislava 811 04 Slovakia

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$a Kvapilova, Katerina $u Faculty of Science, Charles University, Albertov 6, Prague, 128 00, Czech Republic. kvapilova@iabio.eu $u Institute of Applied Biotechnologies a.s, Služeb 4, Prague, 108 00, Czech Republic. kvapilova@iabio.eu

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$a Validated WGS and WES protocols proved saliva-derived gDNA as an equivalent to blood-derived gDNA for clinical and population genomic analyses / $c K. Kvapilova, P. Misenko, J. Radvanszky, O. Brzon, J. Budis, J. Gazdarica, O. Pos, M. Korabecna, M. Kasny, T. Szemes, P. Kvapil, J. Paces, Z. Kozmik

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$a BACKGROUND: Whole exome sequencing (WES) and whole genome sequencing (WGS) have become standard methods in human clinical diagnostics as well as in population genomics (POPGEN). Blood-derived genomic DNA (gDNA) is routinely used in the clinical environment. Conversely, many POPGEN studies and commercial tests benefit from easy saliva sampling. Here, we evaluated the quality of variant call sets and the level of genotype concordance of single nucleotide variants (SNVs) and small insertions and deletions (indels) for WES and WGS using paired blood- and saliva-derived gDNA isolates employing genomic reference-based validated protocols. METHODS: The genomic reference standard Coriell NA12878 was repeatedly analyzed using optimized WES and WGS protocols, and data calls were compared with the truth dataset published by the Genome in a Bottle Consortium. gDNA was extracted from the paired blood and saliva samples of 10 participants and processed using the same protocols. A comparison of paired blood-saliva call sets was performed in the context of WGS and WES genomic reference-based technical validation results. RESULTS: The quality pattern of called variants obtained from genomic-reference-based technical replicates correlates with data calls of paired blood-saliva-derived samples in all levels of tested examinations despite a higher rate of non-human contamination found in the saliva samples. The F1 score of 10 blood-to-saliva-derived comparisons ranged between 0.8030-0.9998 for SNVs and between 0.8883-0.9991 for small-indels in the case of the WGS protocol, and between 0.8643-0.999 for SNVs and between 0.7781-1.000 for small-indels in the case of the WES protocol. CONCLUSION: Saliva may be considered an equivalent material to blood for genetic analysis for both WGS and WES under strict protocol conditions. The accuracy of sequencing metrics and variant-detection accuracy is not affected by choosing saliva as the gDNA source instead of blood but much more significantly by the genomic context, variant types, and the sequencing technology used.

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