Beyond the exome: What’s next in diagnostic testing for Mendelian conditions

The American Journal of Human Genetics, Journal Year: 2023, Volume and Issue: 110(8), P. 1229 - 1248

Published: Aug. 1, 2023

Language: Английский

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Applications of long-read sequencing to Mendelian genetics

Genome Medicine, Journal Year: 2023, Volume and Issue: 15(1)

Published: June 14, 2023

Advances in clinical genetic testing, including the introduction of exome sequencing, have uncovered molecular etiology for many rare and previously unsolved disorders, yet more than half individuals with a suspected disorder remain after complete evaluation. A precise diagnosis may guide treatment plans, allow families to make informed care decisions, permit participate N-of-1 trials; thus, there is high interest developing new tools techniques increase solve rate. Long-read sequencing (LRS) promising technology both increasing rate decreasing amount time required diagnosis. Here, we summarize current LRS technologies, give examples how they been used evaluate complex variation identify missing variants, discuss future applications LRS. As costs continue decrease, will find additional utility space fundamentally changing pathological variants are discovered eventually acting as single-data source that can be interrogated multiple times service.

Language: Английский

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Nanopore DNA sequencing technologies and their applications towards single-molecule proteomics

Nature Chemistry, Journal Year: 2024, Volume and Issue: 16(3), P. 314 - 334

Published: March 1, 2024

Language: Английский

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Sequencing accuracy and systematic errors of nanopore direct RNA sequencing

BMC Genomics, Journal Year: 2024, Volume and Issue: 25(1)

Published: May 28, 2024

Abstract Background Direct RNA sequencing (dRNA-seq) on the Oxford Nanopore Technologies (ONT) platforms can produce reads covering up to full-length gene transcripts, while containing decipherable information about base modifications and poly-A tail lengths. Although many published studies have been expanding potential of dRNA-seq, its accuracy error patterns remain understudied. Results We present first comprehensive evaluation characterisation systematic errors in dRNA-seq data from diverse organisms synthetic vitro transcribed RNAs. found that for kits SQK-RNA001 SQK-RNA002, median read ranged 87% 92% across species, deletions significantly outnumbered mismatches insertions. Due their high abundance transcriptome, heteropolymers short homopolymers were major contributors overall errors. also observed biases all species at levels single nucleotides motifs. In general, cytosine/uracil-rich regions more likely be erroneous than guanines adenines. By examining raw signal data, we identified underlying signal-level features potentially associated with dependency sequence contexts. While quality scores used approximate rates levels, failure detect DNA adapters may a source loss. comparing distinct basecallers, reason some are attributable insufficiency rather algorithmic (basecalling) artefacts. Lastly, generated using latest SQK-RNA004 kit released end 2023 although increased, largely identical compared previous kits. Conclusions As investigation errors, this study offers overview reproducible datasets, identifies insufficiency, lays foundation correction methods.

Language: Английский

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Single-molecule epitranscriptomic analysis of full-length HIV-1 RNAs reveals functional roles of site-specific m6As

Nature Microbiology, Journal Year: 2024, Volume and Issue: 9(5), P. 1340 - 1355

Published: April 11, 2024

Abstract Although the significance of chemical modifications on RNA is acknowledged, evolutionary benefits and specific roles in human immunodeficiency virus (HIV-1) replication remain elusive. Most studies have provided only population-averaged values for fragmented RNAs at low resolution relied indirect analyses phenotypic effects by perturbing host effectors. Here we analysed HIV-1 full-length, single level nucleotide using direct sequencing methods. Our data reveal an unexpectedly simple modification landscape, highlighting three predominant N 6 -methyladenosine (m A) near 3′ end. More densely installed spliced viral messenger than genomic RNAs, these m As play a crucial role maintaining normal levels splicing translation. generates diverse subspecies with distinct A ensembles, multiple its provides additional stability resilience to replication, suggesting unexplored RNA-level strategy.

Language: Английский

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Profiling the epigenome using long-read sequencing

Nature Genetics, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 8, 2025

Language: Английский

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De novo basecalling of RNA modifications at single molecule and nucleotide resolution

Genome biology, Journal Year: 2025, Volume and Issue: 26(1)

Published: Feb. 25, 2025

Abstract RNA modifications influence function and fate, but detecting them in individual molecules remains challenging for most modifications. Here we present a novel methodology to generate training sets build modification-aware basecalling models. Using this approach, develop the m 6 ABasecaller , model that predicts A from raw nanopore signals. We validate its accuracy vitro vivo, revealing stable modification stoichiometry across isoforms, co-occurrence within molecules, A-dependent effects on poly(A) tails. Finally, demonstrate our method generalizes other DNA modifications, paving path towards future efforts

Language: Английский

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Beyond assembly: the increasing flexibility of single-molecule sequencing technology

Nature Reviews Genetics, Journal Year: 2023, Volume and Issue: 24(9), P. 627 - 641

Published: May 9, 2023

Language: Английский

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Current concepts, advances, and challenges in deciphering the human microbiota with metatranscriptomics

Trends in Genetics, Journal Year: 2023, Volume and Issue: 39(9), P. 686 - 702

Published: June 25, 2023

Metatranscriptomics refers to the analysis of collective microbial transcriptome a sample. Its increased utilization for characterization human-associated communities has enabled discovery many disease-state related activities. Here, we review principles metatranscriptomics-based samples. We describe strengths and weaknesses popular sample preparation, sequencing, bioinformatics approaches summarize strategies their use. then discuss how have recently been examined may change. conclude that metatranscriptomics insights into human microbiotas under health disease not only expanded our knowledge on health, but also opened avenues rational antimicrobial drug use management.

Language: Английский

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Transfer learning enables identification of multiple types of RNA modifications using nanopore direct RNA sequencing

Nature Communications, Journal Year: 2024, Volume and Issue: 15(1)

Published: May 14, 2024

Abstract Nanopore direct RNA sequencing (DRS) has emerged as a powerful tool for modification identification. However, concurrently detecting multiple types of modifications in single DRS sample remains challenge. Here, we develop TandemMod, transferable deep learning framework capable data. To train high-performance TandemMod models, generate vitro epitranscriptome datasets from cDNA libraries, containing thousands transcripts labeled with various modifications. We validate the performance on both and vivo human cell lines, confirming its high accuracy profiling m 6 A 5 C sites. Furthermore, perform transfer identifying other such 7 G, Ψ, inosine, significantly reducing training data size running time without compromising performance. Finally, apply to identify 3 rice grown different environments, demonstrating applicability across species conditions. In summary, provide resource ground-truth labels that can serve benchmark nanopore-based identification methods, diverse using sample.

Language: Английский

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