| 1. |
- Ameur, Adam, et al.
(författare)
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De Novo Assembly of Two Swedish Genomes Reveals Missing Segments from the Human GRCh38 Reference and Improves Variant Calling of Population-Scale Sequencing Data
- 2018
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Ingår i: Genes. - : MDPI AG. - 2073-4425 .- 2073-4425. ; 9:10
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 2. |
- Ameur, Adam, et al.
(författare)
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SweGen : a whole-genome data resource of genetic variability in a cross-section of the Swedish population
- 2017
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Ingår i: European Journal of Human Genetics. - : NATURE PUBLISHING GROUP. - 1018-4813 .- 1476-5438. ; 25:11, s. 1253-1260
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Tidskriftsartikel (refereegranskat)abstract
- Here we describe the SweGen data set, a comprehensive map of genetic variation in the Swedish population. These data represent a basic resource for clinical genetics laboratories as well as for sequencing-based association studies by providing information on genetic variant frequencies in a cohort that is well matched to national patient cohorts. To select samples for this study, we first examined the genetic structure of the Swedish population using high-density SNP-array data from a nation-wide cohort of over 10 000 Swedish-born individuals included in the Swedish Twin Registry. A total of 1000 individuals, reflecting a cross-section of the population and capturing the main genetic structure, were selected for whole-genome sequencing. Analysis pipelines were developed for automated alignment, variant calling and quality control of the sequencing data. This resulted in a genome-wide collection of aggregated variant frequencies in the Swedish population that we have made available to the scientific community through the website https://swefreq.nbis.se. A total of 29.2 million single-nucleotide variants and 3.8 million indels were detected in the 1000 samples, with 9.9 million of these variants not present in current databases. Each sample contributed with an average of 7199 individual-specific variants. In addition, an average of 8645 larger structural variants (SVs) were detected per individual, and we demonstrate that the population frequencies of these SVs can be used for efficient filtering analyses. Finally, our results show that the genetic diversity within Sweden is substantial compared with the diversity among continental European populations, underscoring the relevance of establishing a local reference data set.
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| 3. |
- Halvardson, Jonatan, et al.
(författare)
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Mutations in HECW2 are associated with intellectual disability and epilepsy
- 2016
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Ingår i: Journal of Medical Genetics. - : BMJ. - 0022-2593 .- 1468-6244. ; 53:10, s. 697-704
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: De novo mutations are a frequent cause of disorders related to brain development. We report the results of screening patients diagnosed with both epilepsy and intellectual disability (ID) using exome sequencing to identify known and new causative de novo mutations relevant to these conditions.METHODS: Exome sequencing was performed on 39 patient-parent trios to identify de novo mutations. Clinical significance of de novo mutations in genes was determined using the American College of Medical Genetics and Genomics standard guidelines for interpretation of coding variants. Variants in genes of unknown clinical significance were further analysed in the context of previous trio sequencing efforts in neurodevelopmental disorders.RESULTS: In 39 patient-parent trios we identified 29 de novo mutations in coding sequence. Analysis of de novo and inherited variants yielded a molecular diagnosis in 11 families (28.2%). In combination with previously published exome sequencing results in neurodevelopmental disorders, our analysis implicates HECW2 as a novel candidate gene in ID and epilepsy.CONCLUSIONS: Our results support the use of exome sequencing as a diagnostic approach for ID and epilepsy, and confirm previous results regarding the importance of de novo mutations in this patient group. The results also highlight the utility of network analysis and comparison to previous large-scale studies as strategies to prioritise candidate genes for further studies. This study adds knowledge to the increasingly growing list of causative and candidate genes in ID and epilepsy and highlights HECW2 as a new candidate gene for neurodevelopmental disorders.
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| 4. |
- Halvardson, Jonatan, 1982-
(författare)
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Sequence based analysis of neurodevelopmental disorders
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In this thesis the main focus is the use of methods and applications of next generation sequencing in order to study three of the most common neurodevelopmental disorders: intellectual disability, epilepsy and schizophrenia. A large fraction of the genes in our genome produce several distinct transcript isoforms through the process of splicing and there is an increasing amount of evidence pinpointing mutations affecting splicing as a mechanism of disease. In Paper I we used exome capture of RNA in combination with sequencing in order to enrich for coding sequences. We show that this approach enables us to detect lowly expressed transcript and splice events that would have been missed in regular RNA sequencing using the same coverage. In Paper II we selectively depleted the different transcripts of Quaking (QKI), a gene previously associated to schizophrenia. Using RNA sequencing we show that the effects of depletion differ between transcripts and that the QKI gene is a potential regulator of the Glial Fibrillary Acidic Protein (GFAP), a gene implicated in several diseases in the central nervous system.De-novo mutations are frequently reported to be causative in neurodevelopmental disorders with a strong genetic component, such as epilepsy and intellectual disability. In Paper III we used exome sequencing in family trios where the child was diagnosed with both intellectual disability and epilepsy, focusing on finding de-novo mutations. We identified several previously unknown disease causing mutations in genes previously known to cause disease and used previously published interaction and mutation data to prioritize novel candidate genes. The most interesting result from this study are the implication of the HECW2 gene as a candidate gene in intellectual disability and epilepsy. In Paper IV we used RNA sequencing of post mortem brain tissue in a large cohort of schizophrenics and controls. In this study we could show that the immune system and more specifically the complement system was dysregulated in a large fraction of patients. Further, using co-expression network we also found some evidence suggesting genes involved in axon development and maintenance.
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| 5. |
- Höijer, Ida, et al.
(författare)
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Detailed analysis of HTT repeat elements in human blood using targeted amplification-free long-read sequencing
- 2018
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Ingår i: Human Mutation. - : Hindawi Limited. - 1059-7794 .- 1098-1004. ; 39:9, s. 1262-1272
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Tidskriftsartikel (refereegranskat)abstract
- 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.
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| 6. |
- Johansson, Martin M., 1976-, et al.
(författare)
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Novel Y-Chromosome Long Non-Coding RNAs Expressed in Human Male CNS During Early Development
- 2019
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Ingår i: Frontiers in Genetics. - : Frontiers Media S.A.. - 1664-8021. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Global microarray gene expression analyses previously demonstrated differences in female and male embryos during neurodevelopment. In particular, before sexual maturation of the gonads, the differences seem to concentrate on the expression of genes encoded on the X- and Y-chromosomes. To investigate genome-wide differences in expression during this early developmental window, we combined high-resolution RNA sequencing with qPCR to analyze brain samples from human embryos during the first trimester of development. Our analysis was tailored for maximum sensitivity to discover Y-chromosome gene expression, but at the same time, it was underpowered to detect X-inactivation escapees. Using this approach, we found that 5 out of 13 expressed gametolog pairs showed unbalanced gene dosage, and as a consequence, a male-biased expression. In addition, we found six novel non-annotated long non-coding RNAs on the Y-chromosome with conserved expression patterns in newborn chimpanzee. The tissue specific and time-restricted expression of these long non-coding RNAs strongly suggests important functions during central nervous system development in human males.
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| 7. |
- Johansson, Martin, 1976-, et al.
(författare)
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Spatial sexual dimorphism of X and Y homolog gene expression in the human central nervous system during early male development
- 2016
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Ingår i: Biology of Sex Differences. - : Springer Science and Business Media LLC. - 2042-6410. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- Background: Renewed attention has been directed to the functions of the Y chromosome in the central nervous system during early human male development, due to the recent proposed involvement in neurodevelopmental diseases. PCDH11Y and NLGN4Y are of special interest because they belong to gene families involved in cell fate determination and formation of dendrites and axon. Methods: We used RNA sequencing, immunocytochemistry and a padlock probing and rolling circle amplification strategy, to distinguish the expression of X and Y homologs in situ in the human brain for the first time. To minimize influence of androgens on the sex differences in the brain, we focused our investigation to human embryos at 8-11 weeks post-gestation. Results: We found that the X- and Y-encoded genes are expressed in specific and heterogeneous cellular sub-populations of both glial and neuronal origins. More importantly, we found differential distribution patterns of X and Y homologs in the male developing central nervous system. Conclusions: This study has visualized the spatial distribution of PCDH11X/Y and NLGN4X/Y in human developing nervous tissue. The observed spatial distribution patterns suggest the existence of an additional layer of complexity in the development of the male CNS.
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| 8. |
- Kuderna, Lukas F. K., et al.
(författare)
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A 3-way hybrid approach to generate a new high-quality chimpanzee reference genome (Pan_tro_3.0)
- 2017
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Ingår i: GigaScience. - : Oxford University Press (OUP). - 2047-217X. ; 6:11, s. 1-6
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Tidskriftsartikel (refereegranskat)abstract
- The chimpanzee is arguably the most important species for the study of human origins. A key resource for these studies is a high-quality reference genome assembly; however, as with most mammalian genomes, the current iteration of the chimpanzee reference genome assembly is highly fragmented. In the current iteration of the chimpanzee reference genome assembly (Pan tro 2.1.4), the sequence is scattered across more then 183 000 contigs, incorporating more than 159 000 gaps, with a genome-wide contig N50 of 51 Kbp. In this work, we produce an extensive and diverse array of sequencing datasets to rapidly assemble a new chimpanzee reference that surpasses previous iterations in bases represented and organized in large scaffolds. To this end, we show substantial improvements over the current release of the chimpanzee genome (Pan tro 2.1.4) by several metrics, such as increased contiguity by > 750% and 300% on contigs and scaffolds, respectively, and closure of 77% of gaps in the Pan tro 2.1.4 assembly gaps spanning > 850 Kbp of the novel coding sequence based on RNASeq data. We further report more than 2700 genes that had putatively erroneous frame-shift predictions to human in Pan tro 2.1.4 and show a substantial increase in the annotation of repetitive elements. We apply a simple 3-way hybrid approach to considerably improve the reference genome assembly for the chimpanzee, providing a valuable resource for the study of human origins. Furthermore, we produce extensive sequencing datasets that are all derived from the same cell line, generating a broad non-human benchmark dataset.
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| 9. |
- Lindholm Carlström, Eva, et al.
(författare)
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Linkage and exome analysis implicate multiple genes in non-syndromic intellectual disability in a large Swedish family
- 2019
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Ingår i: BMC Medical Genomics. - : Springer Science and Business Media LLC. - 1755-8794. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundNon-syndromic intellectual disability is genetically heterogeneous with dominant, recessive and complex forms of inheritance. We have performed detailed genetic studies in a large multi-generational Swedish family, including several members diagnosed with non-syndromic intellectual disability. Linkage analysis was performed on 22 family members, nine affected with mild to moderate intellectual disability and 13 unaffected family members.Methods Family members were analyzed with Affymetrix Genome-Wide Human SNP Array 6.0 and the genetic data was used to detect copy number variation and to perform genome wide linkage analysis with the SNP High Throughput Linkage analysis system and the Merlin software. For the exome sequencing, the samples were prepared using the Sure Select Human All Exon Kit (Agilent Technologies, Santa Clara, CA, USA) and sequenced using the Ion Proton (TM) System. Validation of identified variants was performed with Sanger sequencing.ResultsThe linkage analysis results indicate that intellectual disability in this family is genetically heterogeneous, with suggestive linkage found on chromosomes 1q31-q41, 4q32-q35, 6p25 and 14q24-q31 (LOD scores of 2.4, simulated p-value of 0.000003 and a simulated genome-wide p-value of 0.06). Exome sequencing was then performed in 14 family members and 7 unrelated individuals from the same region. The analysis of coding variation revealed a pathogenic and candidate variants in different branches of the family. In three patients we find a known homozygous pathogenic mutation in the Homo sapiens solute carrier family 17 member 5 (SLC17A5), causing Salla disease. We also identify a deletion overlapping KDM3B and a duplication overlapping MAP3K4 and AGPAT4, both overlapping variants previously reported in developmental disorders.Conclusions DNA samples from the large family analyzed in this study were initially collected based on a hypothesis that affected members shared a major genetic risk factor. Our results show that a complex phenotype such as mild intellectual disability in large families from genetically isolated populations may show considerable genetic heterogeneity.
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| 10. |
- Magnusson, Patrik K. E., et al.
(författare)
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One CNV Discordance in NRXN1 Observed Upon Genome-wide Screening in 38 Pairs of Adult Healthy Monozygotic Twins
- 2016
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Ingår i: Twin Research and Human Genetics. - : Cambridge University Press (CUP). - 1832-4274 .- 1839-2628. ; 19:2, s. 97-103
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Tidskriftsartikel (refereegranskat)abstract
- Monozygotic (MZ) twins stem from the same single fertilized egg and therefore share all their inherited genetic variation. This is one of the unequivocal facts on which genetic epidemiology and twin studies are based. To what extent this also implies that MZ twins share genotypes in adult tissues is not precisely established, but a common pragmatic assumption is that MZ twins are 100% genetically identical also in adult tissues. During the past decade, this view has been challenged by several reports, with observations of differences in post-zygotic copy number variations (CNVs) between members of the same MZ pair. In this study, we performed a systematic search for differences of CNVs within 38 adult MZ pairs who had been misclassified as dizygotic (DZ) twins by questionnaire-based assessment. Initial scoring by PennCNV suggested a total of 967 CNV discor dances. The within-pair correlation in number of CNVs detected was strongly dependent on confidence score filtering and reached a plateau of r = 0.8 when restricting to CNVs detected with confidence score larger than 50. The top-ranked discordances were subsequently selected for validation by quantitative polymerase chain reaction (qPCR), from which one single similar to 120kb deletion in NRXN1 on chromosome 2 (bp 51017111-51136802) was validated. Despite involving an exon, no sign of cognitive/mental consequences was apparent in the affected twin pair, potentially reflecting limited or lack of expression of the transcripts containing this exon in nerve/brain.
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