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Ameur, A. (2019). Goodbye reference, hello genome graphs. Nature Biotechnology, 37(8), 866-868
Open this publication in new window or tab >>Goodbye reference, hello genome graphs
2019 (English)In: Nature Biotechnology, ISSN 1087-0156, E-ISSN 1546-1696, Vol. 37, no 8, p. 866-868Article in journal, Editorial material (Other academic) Published
Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2019
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-394149 (URN)10.1038/s41587-019-0199-7 (DOI)000482876100017 ()31375808 (PubMedID)
Available from: 2019-10-04 Created: 2019-10-04 Last updated: 2019-10-04Bibliographically approved
Cui, T., Enroth, S., Ameur, A., Gustavsson, I., Lindquist, D. & Gyllensten, U. (2019). Invasive cervical tumors with high and low HPV titer represent molecular subgroups with different disease etiology. Carcinogenesis, 40(2), 269-278
Open this publication in new window or tab >>Invasive cervical tumors with high and low HPV titer represent molecular subgroups with different disease etiology
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2019 (English)In: Carcinogenesis, ISSN 0143-3334, E-ISSN 1460-2180, Vol. 40, no 2, p. 269-278Article in journal (Refereed) Published
Abstract [en]

Invasive cervical cancer (ICC) with very low titer of high-risk human papillomavirus (HPV) has worse clinical outcome than cases with high titer, indicating a difference in molecular etiology. Fresh-frozen ICC tumors (n = 49) were classified into high- and low-HPV-titer cases using real-time PCR-based HPV genotyping. The mutation spectra were studied using the AmpliSeq Comprehensive Cancer Panel and the expression profiles using total RNA sequencing, and the results were validated using the AmpliSeq Transcriptome assay. HPV DNA genotyping and RNA sequencing showed that 16.6% of ICC tumors contained very low levels of HPV DNA and HPV transcripts. Tumors with low HPV levels had more mutations with a high allele frequency and fewer mutations with low allele frequency relative to tumors with high HPV titer. A number of genes showed significant expression differences between HPV titer groups, including genes with somatic mutations. Gene ontology and pathway analyses implicated the enrichment of genes involved in DNA replication, cell cycle control and extracellular matrix in tumors with low HPV titer. The results indicate that in low titer tumors, HPVs act as trigger of cancer development whereas somatic mutations are clonally selected and become drivers of the tumor development process. In contrast, in tumors with high HPV titer the expression of HPV oncoproteins plays a major role in tumor development and the many low frequency somatic mutations represent passengers. This putative subdivision of invasive cervical tumors may explain the higher radiosensitivity of ICC tumors with high HPV titer and thereby have consequences for clinical management.

National Category
Cell and Molecular Biology Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-388612 (URN)10.1093/carcin/bgy183 (DOI)000472792800012 ()30596972 (PubMedID)
Funder
Swedish Cancer Society, CF23560Swedish Research Council, 2012-2884Swedish Society for Medical Research (SSMF)Knut and Alice Wallenberg Foundation, KAW2011.0205
Note

De 2 första författarna delar förstaförfattarskapet.

Available from: 2019-07-02 Created: 2019-07-02 Last updated: 2019-08-23Bibliographically approved
Ameur, A., Kloosterman, W. P. & Hestand, M. S. (2019). Single-Molecule Sequencing: Towards Clinical Applications. Trends in Biotechnology, 37(1), 72-85
Open this publication in new window or tab >>Single-Molecule Sequencing: Towards Clinical Applications
2019 (English)In: Trends in Biotechnology, ISSN 0167-7799, E-ISSN 1879-3096, Vol. 37, no 1, p. 72-85Article, review/survey (Refereed) Published
Abstract [en]

In the past several years, single-molecule sequencing platforms, such as those by Pacific Biosciences and Oxford Nanopore Technologies, have become available to researchers and are currently being tested for clinical applications. They offer exceptionally long reads that permit direct sequencing through regions of the genome inaccessible or difficult to analyze by short-read platforms. This includes disease-causing long repetitive elements, extreme GC content regions, and complex gene loci. Similarly, these platforms enable structural variation characterization at previously unparalleled resolution and direct detection of epigenetic marks in native DNA. Here, we review how these technologies are opening up new clinical avenues that are being applied to pathogenic microorganisms and viruses, constitutional disorders, pharmaco-genomics, cancer, and more.

National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-373914 (URN)10.1016/j.tibtech.2018.07.013 (DOI)000453919500009 ()30115375 (PubMedID)
Available from: 2019-01-17 Created: 2019-01-17 Last updated: 2019-01-17Bibliographically approved
Hestand, M. S. & Ameur, A. (2019). The Versatility of SMRT Sequencing. Genes, 10(1), Article ID 24.
Open this publication in new window or tab >>The Versatility of SMRT Sequencing
2019 (English)In: Genes, ISSN 2073-4425, E-ISSN 2073-4425, Vol. 10, no 1, article id 24Article in journal, Editorial material (Other academic) Published
Place, publisher, year, edition, pages
MDPI, 2019
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-379426 (URN)10.3390/genes10010024 (DOI)000459743800024 ()30621217 (PubMedID)
Available from: 2019-03-18 Created: 2019-03-18 Last updated: 2019-03-18Bibliographically approved
Stattin, E.-L., Johansson, J., Gudmundsson, S., Ameur, A., Lundberg, S., Bondeson, M.-L. & Wilbe, M. (2018). A novel ECEL1 mutation expands the phenotype of distal arthrogryposis multiplex congenita type 5D to include pretibial vertical skin crease. American Journal of Medical Genetics. Part A, 176(6), 1405-1410
Open this publication in new window or tab >>A novel ECEL1 mutation expands the phenotype of distal arthrogryposis multiplex congenita type 5D to include pretibial vertical skin crease
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2018 (English)In: American Journal of Medical Genetics. Part A, ISSN 1552-4825, E-ISSN 1552-4833, Vol. 176, no 6, p. 1405-1410Article in journal (Refereed) Published
Abstract [en]

Arthrogryposis multiplex congenita (AMC) is a heterogeneous disorder characterized by multiple joint contractures often in association with other congenital abnormalities. Pretibial linear vertical creases are a rare finding associated with arthrogryposis, and the etiology of the specific condition is unknown. We aimed to genetically and clinically characterize a boy from a consanguineous family, presenting with AMC and pretibial vertical linear creases on the shins. Whole exome sequencing and variant analysis revealed homozygous novel missense variants of ECEL1 (c.1163T > C, p.Leu388Pro, NM_004826) and MUSK (c.2572C > T, p.Arg858Cys, NM_005592). Both variants are predicted to have deleterious effects on the protein function, with amino acid positions highly conserved among species. The variants segregated in the family, with healthy mother, father, and sister being heterozygous carriers and the index patient being homozygous for both mutations. We report on a unique patient with a novel ECEL1 homozygous mutation, expanding the phenotypic spectrum of Distal AMC Type 5D to include vertical linear skin creases. The homozygous mutation in MUSK is of unknown clinical significance. MUSK mutations have previously shown to cause congenital myasthenic syndrome, a neuromuscular disorder with defects in the neuromuscular junction.

Place, publisher, year, edition, pages
WILEY, 2018
Keywords
arthrogryposis multiplex congenital, congenital myasthenic syndrome, ECEL1, vertical linear skin creases, whole exome sequencing
National Category
Medical Genetics
Identifiers
urn:nbn:se:uu:diva-364512 (URN)10.1002/ajmg.a.38691 (DOI)000434040600017 ()29663639 (PubMedID)
Available from: 2018-11-05 Created: 2018-11-05 Last updated: 2018-11-05Bibliographically approved
Ameur, A., Che, H., Martin, M., Bunikis, I., Dahlberg, J., Höijer, I., . . . Gyllensten, U. B. (2018). De Novo Assembly of Two Swedish Genomes Reveals Missing Segments from the Human GRCh38 Reference and Improves Variant Calling of Population-Scale Sequencing Data. Genes, 9(10), Article ID 486.
Open this publication in new window or tab >>De Novo Assembly of Two Swedish Genomes Reveals Missing Segments from the Human GRCh38 Reference and Improves Variant Calling of Population-Scale Sequencing Data
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2018 (English)In: Genes, ISSN 2073-4425, E-ISSN 2073-4425, Vol. 9, no 10, article id 486Article in journal (Refereed) Published
Abstract [en]

The current human reference sequence (GRCh38) is a foundation for large-scale sequencing projects. However, recent studies have suggested that GRCh38 may be incomplete and give a suboptimal representation of specific population groups. Here, we performed a de novo assembly of two Swedish genomes that revealed over 10 Mb of sequences absent from the human GRCh38 reference in each individual. Around 6 Mb of these novel sequences (NS) are shared with a Chinese personal genome. The NS are highly repetitive, have an elevated GC-content, and are primarily located in centromeric or telomeric regions. Up to 1 Mb of NS can be assigned to chromosome Y, and large segments are also missing from GRCh38 at chromosomes 14, 17, and 21. Inclusion of NS into the GRCh38 reference radically improves the alignment and variant calling from short-read whole-genome sequencing data at several genomic loci. A re-analysis of a Swedish population-scale sequencing project yields > 75,000 putative novel single nucleotide variants (SNVs) and removes > 10,000 false positive SNV calls per individual, some of which are located in protein coding regions. Our results highlight that the GRCh38 reference is not yet complete and demonstrate that personal genome assemblies from local populations can improve the analysis of short-read whole-genome sequencing data.

Keywords
de novo assembly, SMRT sequencing, GRCh38, human reference genome, human whole-genome sequencing, population sequencing, Swedish population
National Category
Genetics
Identifiers
urn:nbn:se:uu:diva-369762 (URN)10.3390/genes9100486 (DOI)000448656700024 ()30304863 (PubMedID)
Funder
Knut and Alice Wallenberg Foundation, 2014.0272Swedish Research Council
Available from: 2018-12-17 Created: 2018-12-17 Last updated: 2018-12-17Bibliographically approved
Höijer, I., Tsai, Y.-C., Clark, T. A., Kotturi, P., Dahl, N., Stattin, E., . . . Ameur, A. (2018). Detailed analysis of HTT repeat elements in human blood using targeted amplification-free long-read sequencing. Human Mutation, 39(9), 1262-1272
Open this publication in new window or tab >>Detailed analysis of HTT repeat elements in human blood using targeted amplification-free long-read sequencing
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2018 (English)In: Human Mutation, ISSN 1059-7794, E-ISSN 1098-1004, Vol. 39, no 9, p. 1262-1272Article in journal (Refereed) Published
Abstract [en]

Amplification of DNA is required as a mandatory step during library preparation in most targeted sequencing protocols. This can be a critical limitation when targeting regions that are highly repetitive or with extreme guanine-cytosine (GC) content, including repeat expansions associated with human disease. Here, we used an amplification-free protocol for targeted enrichment utilizing the CRISPR/Cas9 system (No-Amp Targeted sequencing) in combination with single molecule, real-time (SMRT) sequencing for studying repeat elements in the huntingtin (HTT) gene, where an expanded CAG repeat is causative for Huntington disease. We also developed a robust data analysis pipeline for repeat element analysis that is independent of alignment of reads to a reference genome. The method was applied to 11 diagnostic blood samples, and for all 22 alleles the resulting CAG repeat count agreed with previous results based on fragment analysis. The amplification-free protocol also allowed for studying somatic variability of repeat elements in our samples, without the interference of PCR stutter. In summary, with No-Amp Targeted sequencing in combination with our analysis pipeline, we could accurately study repeat elements that are difficult to investigate using PCR-based methods.

Keywords
amplification-free sequencing, HTT, Huntington disease, No-Amp Targeted sequencing, repeat expansion, SMRT sequencing, somatic mosaicism, targeted enrichment, targeted sequencing
National Category
Medical Genetics
Identifiers
urn:nbn:se:uu:diva-364189 (URN)10.1002/humu.23580 (DOI)000443229000010 ()29932473 (PubMedID)
Available from: 2018-11-07 Created: 2018-11-07 Last updated: 2018-11-16Bibliographically approved
Zaghlool, A., Ameur, A., Wu, C., Westholm, J. O., Niazi, A., Manivannan, M., . . . Feuk, L. (2018). Expression profiling and in situ screening of circular RNAs in human tissues. Scientific Reports, 8, Article ID 16953.
Open this publication in new window or tab >>Expression profiling and in situ screening of circular RNAs in human tissues
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2018 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, article id 16953Article in journal (Refereed) Published
Abstract [en]

Circular RNAs (circRNAs) were recently discovered as a class of widely expressed noncoding RNA and have been implicated in regulation of gene expression. However, the function of the majority of circRNAs remains unknown. Studies of circRNAs have been hampered by a lack of essential approaches for detection, quantification and visualization. We therefore developed a target-enrichment sequencing method suitable for screening of circRNAs and their linear counterparts in large number of samples. We also applied padlock probes and in situ sequencing to visualize and determine circRNA localization in human brain tissue at subcellular levels. We measured circRNA abundance across different human samples and tissues. Our results highlight the potential of this RNA class to act as a specific diagnostic marker in blood and serum, by detection of circRNAs from genes exclusively expressed in the brain. The powerful and scalable tools we present will enable studies of circRNA function and facilitate screening of circRNA as diagnostic biomarkers.

Place, publisher, year, edition, pages
NATURE PUBLISHING GROUP, 2018
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-371872 (URN)10.1038/s41598-018-35001-6 (DOI)000450280500013 ()30446675 (PubMedID)
Funder
Swedish Research Council, 2012-4530Swedish Research Council, 2017-01861EU, European Research Council, 282330Knut and Alice Wallenberg Foundation
Available from: 2019-01-07 Created: 2019-01-07 Last updated: 2019-01-07Bibliographically approved
Ardui, S., Ameur, A., Vermeesch, J. R. & Hestand, M. S. (2018). Single molecule real-time (SMRT) sequencing comes of age: applications and utilities for medical diagnostics. Nucleic Acids Research, 46(5), 2159-2168
Open this publication in new window or tab >>Single molecule real-time (SMRT) sequencing comes of age: applications and utilities for medical diagnostics
2018 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 46, no 5, p. 2159-2168Article in journal (Refereed) Published
Abstract [en]

Short read massive parallel sequencing has emerged as a standard diagnostic tool in the medical setting. However, short read technologies have inherent limitations such as GC bias, difficulties mapping to repetitive elements, trouble discriminating paralogous sequences, and difficulties in phasing alleles. Long read single molecule sequencers resolve these obstacles. Moreover, they offer higher consensus accuracies and can detect epigenetic modifications from native DNA. The first commercially available long read single molecule platform was the RS system based on PacBio's single molecule realtime (SMRT) sequencing technology, which has since evolved into their RSII and Sequel systems. Here we capsulize how SMRT sequencing is revolutionizing constitutional, reproductive, cancer, microbial and viral genetic testing.

Place, publisher, year, edition, pages
OXFORD UNIV PRESS, 2018
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-351426 (URN)10.1093/nar/gky066 (DOI)000427677100008 ()29401301 (PubMedID)
Available from: 2018-06-01 Created: 2018-06-01 Last updated: 2018-06-01Bibliographically approved
Lode, L., Ameur, A., Coste, T., Menard, A., Richebourg, S., Gaillard, J.-B., . . . Soussi, T. (2018). Single-molecule DNA sequencing of acute myeloid leukemia and myelodysplastic syndromes with multiple TP53 alterations [Letter to the editor]. Haematologica, 103(1), Article ID e13.
Open this publication in new window or tab >>Single-molecule DNA sequencing of acute myeloid leukemia and myelodysplastic syndromes with multiple TP53 alterations
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2018 (English)In: Haematologica, ISSN 0390-6078, E-ISSN 1592-8721, Vol. 103, no 1, article id e13Article in journal, Letter (Other academic) Published
Place, publisher, year, edition, pages
FERRATA STORTI FOUNDATION, 2018
National Category
Hematology
Identifiers
urn:nbn:se:uu:diva-386358 (URN)10.3324/haematol.2017.176719 (DOI)000418944500004 ()29079597 (PubMedID)
Available from: 2019-06-19 Created: 2019-06-19 Last updated: 2019-06-19Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0001-6085-6749

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