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Sci Rep. 2018 Mar 28;8(1):5307. doi: 10.1038/s41598-018-23563-4.

Sample-Index Misassignment Impacts Tumour Exome Sequencing.

Vodák D1,2,3, Lorenz S2,3,4, Nakken S2,3, Aasheim LB2, Holte H2,5,6, Bai B6,7, Myklebost O2,3,8, Meza-Zepeda LA2,3,9, Hovig E10,11,12,13.

Author information

1
Institute of Clinical Medicine, Faculty of Medicine, University of Oslo, Oslo, Norway.
2
Norwegian Cancer Genomics Consortium, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway.
3
Department of Tumor Biology, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway.
4
Department of Core Facilities, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway.
5
Department of Oncology, Cancer Clinic, Oslo University Hospital, Oslo, Norway.
6
Centre for Cancer Biomedicine, University of Oslo, Oslo, Norway.
7
Department of Cancer Immunology, Institute for Cancer Research, Oslo University Hospital, Oslo, Norway.
8
Department of Clinical Science, University of Bergen, Bergen, Norway.
9
Genomics Core Facility, Department of Core Facilities, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway.
10
Norwegian Cancer Genomics Consortium, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway. ehovig@ifi.uio.no.
11
Department of Tumor Biology, Institute for Cancer Research, The Norwegian Radium Hospital/Oslo University Hospital, Oslo, Norway. ehovig@ifi.uio.no.
12
Department of Cancer Genetics and Informatics, Oslo University Hospital, Oslo, Norway. ehovig@ifi.uio.no.
13
Department of Informatics, University of Oslo, Oslo, Norway. ehovig@ifi.uio.no.

Abstract

Sample pooling enabled by dedicated indexes is a common strategy for cost-effective and robust high-throughput sequencing. Index misassignment leading to mutual contamination between pooled samples has however been described as a general problem of the latest Illumina sequencing instruments utilizing exclusion amplification. Using real-life data from multiple tumour sequencing projects, we demonstrate that index misassignment can induce artefactual variant calls closely resembling true, high-quality somatic variants. These artefactual calls potentially impact cancer applications utilizing low allelic frequencies, such as in clonal analysis of tumours. We discuss the available countermeasures with an emphasis on improved library indexing methods, and provide software that can assist in the identification of variants that may be consequences of index misassignment.

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