Long-Read DNA Sequencing: Recent Advances and Remaining Challenges

Annual Review of Genomics and Human Genetics, Год журнала: 2023, Номер 24(1), С. 109 - 132

Опубликована: Апрель 19, 2023

DNA sequencing has revolutionized medicine over recent decades. However, analysis of large structural variation and repetitive DNA, a hallmark human genomes, been limited by short-read technology, with read lengths 100-300 bp. Long-read (LRS) permits routine fragments tens to hundreds kilobase pairs in size, using both real-time synthesis nanopore-based direct electronic sequencing. LRS haplotypic phasing genomes enabled the discovery characterization rare pathogenic variants repeat expansions. It also recently assembly complete, gapless genome that includes previously intractable regions, such as highly centromeres homologous acrocentric short arms. With addition protocols for targeted enrichment, epigenetic modification detection, long-range chromatin profiling, promises launch new era understanding genetic diversity mutations populations.

Язык: Английский

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Sequencing and characterizing short tandem repeats in the human genome

Nature Reviews Genetics, Год журнала: 2024, Номер 25(7), С. 460 - 475

Опубликована: Фев. 16, 2024

Язык: Английский

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Sequence composition changes in short tandem repeats: heterogeneity, detection, mechanisms and clinical implications

Nature Reviews Genetics, Год журнала: 2024, Номер 25(7), С. 476 - 499

Опубликована: Март 11, 2024

Язык: Английский

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Long read sequencing on its way to the routine diagnostics of genetic diseases

Frontiers in Genetics, Год журнала: 2024, Номер 15

Опубликована: Март 6, 2024

The clinical application of technological progress in the identification DNA alterations has always led to improvements diagnostic yields genetic medicine. At chromosome side, from cytogenetic techniques evaluating number and gross structural defects genomic microarrays detecting cryptic copy variants, at molecular level, Sanger method studying nucleotide sequence single genes high-throughput next-generation sequencing (NGS) technologies, resolution sensitivity progressively increased expanding considerably range detectable anomalies alongside Mendelian disorders with known causes. However, particular regions (i.e., repetitive GC-rich sequences) are inefficiently analyzed by standard tests, still relying on laborious, time-consuming low-sensitive approaches southern-blot for repeat expansion or long-PCR highly homologous pseudogenes), accounting least part patients undiagnosed disorders. Third generation sequencing, generating long reads improved mappability, is more suitable detection hardly accessible regions. Although recently implemented not yet clinically available, read (LRS) technologies have already shown their potential medicine research that might greatly impact yield reporting times, through translation settings. main investigated LRS concerns variants expansions, probably because evolved as rapidly those dedicated (SNV) identification: gold analyses karyotyping balanced unbalanced rearrangements, respectively, southern blot repeat-primed PCR amplification sizing expanded alleles, impaired limited been significantly advent NGS. Nevertheless, recently, accuracy provided latest product releases, tested also SNV detection, especially pseudogenes haplotype reconstruction assess parental origin alleles de novo pathogenic variants. We provide a review relevant recent scientific papers exploring diagnosis diseases its future applications routine testing.

Язык: Английский

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APOBEC3A deaminates CTG hairpin loops to promote fragility and instability of expanded CAG/CTG repeats

Proceedings of the National Academy of Sciences, Год журнала: 2025, Номер 122(2)

Опубликована: Янв. 7, 2025

CAG/CTG repeats are prone to expansion, causing several inherited human diseases. The initiating sources of DNA damage which lead inaccurate repair the repeat tract cause expansions not fully understood. Expansion-prone actively transcribed and forming stable R-loops with hairpin structures on displaced single-stranded (S-loops). We previously determined that by Saccharomyces cerevisiae cytosine deaminase, Fcy1, was required for both fragility instability tracts engaged in R-loops. To determine whether this mechanism is more universal, we expressed cytidine deaminases APOBEC3A (A3A), APOBEC3B (A3B), or activation-induced deaminase (AID) our yeast system. show mutagenic activity Apolipoprotein B messenger RNA-editing enzyme, catalytic polypeptides causes instability, A3A having greatest effect followed A3B least from AID. A3A-induced exacerbated enrichment at site. A3B-induced dependent MutLγ nuclease a lesser extent, base excision factors. Deaminase assays substrates containing CTG GTC triplet sequences revealed prefers cytidines within loop, bulges stem alter preferred locations. Analysis RNA expression levels cortex samples brain tissue exhibits its elevated Huntington’s disease (HD) patient samples. These results implicate deamination as potential source HD other expansion disorders.

Язык: Английский

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Recurrent repeat expansions in human cancer genomes

Nature, Год журнала: 2022, Номер 613(7942), С. 96 - 102

Опубликована: Дек. 14, 2022

Abstract Expansion of a single repetitive DNA sequence, termed tandem repeat (TR), is known to cause more than 50 diseases 1,2 . However, expansions are often not explored beyond neurological and neurodegenerative disorders. In some cancers, mutations accumulate in short tracts TRs, phenomenon microsatellite instability; however, larger have been systematically analysed cancer 3–8 Here we identified TR 2,622 genomes spanning 29 types. seven types, found 160 recurrent (rREs), most which (155/160) were subtype specific. We that rREs non-uniformly distributed the genome with enrichment near candidate cis -regulatory elements, suggesting potential role gene regulation. One rRE, GAAA-repeat expansion, located regulatory element first intron UGT2B7 was detected 34% renal cell carcinoma samples validated by long-read sequencing. Moreover, preliminary experiments, treating cells harbour this rRE GAAA-targeting molecule led dose-dependent decrease proliferation. Overall, our results suggest may be an important but unexplored source genetic variation human cancer, provide comprehensive catalogue for further study.

Язык: Английский

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Rapid and comprehensive diagnostic method for repeat expansion diseases using nanopore sequencing

npj Genomic Medicine, Год журнала: 2022, Номер 7(1)

Опубликована: Окт. 26, 2022

We developed a diagnostic method for repeat expansion diseases using long-read sequencer to improve currently available, low throughput methods. employed the real-time target enrichment system of nanopore GridION adaptive sampling option, in which software-based assignment is available without prior sample enrichment, and built an analysis pipeline that prioritized disease-causing loci. Twenty-two patients with various neurological neuromuscular diseases, including 12 genetically diagnosed 10 manifesting cerebellar ataxia, but genetic diagnosis, were analyzed. first sequenced molecularly accurately confirmed expanded repeats all uniform depth coverage across Next, we applied our conventional undiagnosed patients. Our corrected inaccurate diagnoses two by method. superior methods terms speed, accuracy, comprehensiveness.

Язык: Английский

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Characterization of genome-wide STR variation in 6487 human genomes

Nature Communications, Год журнала: 2023, Номер 14(1)

Опубликована: Апрель 12, 2023

Short tandem repeats (STRs) are abundant and highly mutagenic in the human genome. Many STR loci have been associated with a range of genetic disorders. However, most population-scale studies on variation humans focused European ancestry cohorts or limited by sequencing depth. Here, we depicted comprehensive map 366,013 polymorphic STRs (pSTRs) constructed from 6487 deeply sequenced genomes, comprising 3983 Chinese samples (~31.5x, NyuWa) 2504 1000 Genomes Project (~33.3x, 1KGP). We found that mutations were affected motif length, chromosome context epigenetic features. identified 3273 1117 pSTRs whose repeat numbers gene expression 3'UTR alternative polyadenylation, respectively. also implemented population analysis, investigated differentiated signatures, genotyped 60 known disease-causing STRs. Overall, this study further extends scale propels our understanding semantics

Язык: Английский

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Detection of alternative DNA structures and its implications for human disease

Molecular Cell, Год журнала: 2023, Номер 83(20), С. 3622 - 3641

Опубликована: Окт. 1, 2023

Язык: Английский

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Genome-wide tandem repeat expansions contribute to schizophrenia risk

Molecular Psychiatry, Год журнала: 2022, Номер 27(9), С. 3692 - 3698

Опубликована: Май 12, 2022

Abstract Tandem repeat expansions (TREs) can cause neurological diseases but their impact in schizophrenia is unclear. Here we analyzed genome sequences of adults with and found that they have a higher burden TREs are near exons rare the general population, compared non-psychiatric controls. These disproportionately at loci known to be associated from genome-wide association studies, individuals clinically-relevant genetic variants other loci, families where multiple schizophrenia. We showed may synaptic functions by disrupting splicing process genes loss-of-function manner. Our findings support involvement polygenic nature

Язык: Английский

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