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Diversity in non-repetitive human sequences not found in the reference genome

Abstract

Genomes usually contain some non-repetitive sequences that are missing from the reference genome and occur only in a population subset. Such non-repetitive, non-reference (NRNR) sequences have remained largely unexplored in terms of their characterization and downstream analyses. Here we describe 3,791 breakpoint-resolved NRNR sequence variants called using PopIns from whole-genome sequence data of 15,219 Icelanders. We found that over 95% of the 244 NRNR sequences that are 200 bp or longer are present in chimpanzees, indicating that they are ancestral. Furthermore, 149 variant loci are in linkage disequilibrium (r2 > 0.8) with a genome-wide association study (GWAS) catalog marker, suggesting disease relevance. Additionally, we report an association (P = 3.8 × 10−8, odds ratio (OR) = 0.92) with myocardial infarction (23,360 cases, 300,771 controls) for a 766-bp NRNR sequence variant. Our results underline the importance of including variation of all complexity levels when searching for variants that associate with disease.

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Figure 1: Workflow for calling non-reference, non-repetitive sequence variants.
Figure 2: Frequency, length and examples of non-reference, non-repetitive sequence variants.
Figure 3: The 766-bp NRNR sequence variant (NRNR1361) affecting an intron of SREBF1 that associates with myocardial infarction.

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NCBI Reference Sequence

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Author information

Authors and Affiliations

Authors

Contributions

B.K., P.M., B.V.H. and K.S. designed the experiments. B.K., P.M., A.G., S.K., D.F.G. and B.V.H. implemented the methodology and analyzed the call set. B.K., A. Helgadottir, H. Holm, P.S., D.F.G. and B.V.H. interpreted the association results. B.K., H. Helgason and G.H.H. analyzed gene expression. Aslaug Jonasdottir, Adalbjorg Jonasdottir, and A.S. performed PCR verification and Sanger sequencing. U.T. oversaw the operations of the genotyping facilities. G.T., I.O., H. Holm and U.T. were responsible for phenotype data acquisition. B.K. prepared tables and figures. B.K., H.J., A. Helgason and B.V.H. wrote the manuscript. All authors reviewed and approved the final manuscript. K.S. supervised the study.

Corresponding authors

Correspondence to Bjarni V Halldorsson or Kari Stefansson.

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Competing interests

B.K., A. Helgadottir, P.M., H.J., H. Helgason, Adalbjorg Jonasdottir, Aslaug Jonasdottir, A.S., A.G., G.H.H., S.K., H. Holm, P.S., U.T., A. Helgason, D.F.G., B.V.H. and K.S. are all employees of deCODE Genetics/Amgen, Inc.

Integrated supplementary information

Supplementary Figure 1 Primer pairs designed for the five categories of NRNR sequence variants for validation by Sanger sequencing.

For the categories “INS > 200” and “DEL > INS”, two or three primer pairs were designed including at least one for each allele. For “INS < 200”, only a single primer pair was designed that may amplify in both alleles. For “Different contig” always three and for “Singleton” always two primer pairs were designed.

Supplementary information

Supplementary Text and Figures

Supplementary Figure 1, Supplementary Tables 2, 7 and 8, and Supplementary Note (PDF 1950 kb)

Supplementary Data 1: NRNR sequences anchored by imputed NRNR markers.

Sequences are given in FASTA format. (TXT 2129 kb)

Supplementary Data 2: NRNR sequences anchored by fixed NRNR markers.

Fixed are those markers that were predicted to have 100% frequency in Iceland. Sequences are given in FASTA format. (TXT 288 kb)

Supplementary Table 1

List of imputed NRNR markers. (XLSX 623 kb)

Supplementary Table 3

Details of Sanger sequencing validation experiments. (XLSX 65 kb)

Supplementary Table 4

List of fixed NRNR markers. (XLSX 48 kb)

Supplementary Table 5

Overlap of NRNR markers with known variants and sequences. (XLSX 358 kb)

Supplementary Table 6

Correlation with the GWAS catalog. (XLSX 84 kb)

Supplementary Table 9

Conversion to GenBank sequences. (XLSX 163 kb)

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Kehr, B., Helgadottir, A., Melsted, P. et al. Diversity in non-repetitive human sequences not found in the reference genome. Nat Genet 49, 588–593 (2017). https://doi.org/10.1038/ng.3801

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