Long read sequencing on its way to the routine diagnostics of genetic diseases
The clinical application of technological progress in the identification of DNA alterations has always led to improvements of diagnostic yields in genetic medicine. At chromosome side, from cytogenetic techniques evaluating number and gross structural defects to genomic microarrays detecting cryptic...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2024-03-01
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| Series: | Frontiers in Genetics |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fgene.2024.1374860/full |
| _version_ | 1827325541022171136 |
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| author | Giulia Olivucci Giulia Olivucci Emanuela Iovino Giovanni Innella Giovanni Innella Daniela Turchetti Daniela Turchetti Tommaso Pippucci Pamela Magini |
| author_facet | Giulia Olivucci Giulia Olivucci Emanuela Iovino Giovanni Innella Giovanni Innella Daniela Turchetti Daniela Turchetti Tommaso Pippucci Pamela Magini |
| author_sort | Giulia Olivucci |
| collection | DOAJ |
| description | The clinical application of technological progress in the identification of DNA alterations has always led to improvements of diagnostic yields in genetic medicine. At chromosome side, from cytogenetic techniques evaluating number and gross structural defects to genomic microarrays detecting cryptic copy number variants, and at molecular level, from Sanger method studying the nucleotide sequence of single genes to the high-throughput next-generation sequencing (NGS) technologies, resolution and sensitivity progressively increased expanding considerably the range of detectable DNA anomalies and alongside of Mendelian disorders with known genetic causes. However, particular genomic regions (i.e., repetitive and GC-rich sequences) are inefficiently analyzed by standard genetic tests, still relying on laborious, time-consuming and low-sensitive approaches (i.e., southern-blot for repeat expansion or long-PCR for genes with highly homologous pseudogenes), accounting for at least part of the patients with undiagnosed genetic disorders. Third generation sequencing, generating long reads with improved mappability, is more suitable for the detection of structural alterations and defects in hardly accessible genomic regions. Although recently implemented and not yet clinically available, long read sequencing (LRS) technologies have already shown their potential in genetic medicine research that might greatly impact on diagnostic yield and reporting times, through their translation to clinical settings. The main investigated LRS application concerns the identification of structural variants and repeat expansions, probably because techniques for their detection have not evolved as rapidly as those dedicated to single nucleotide variants (SNV) identification: gold standard analyses are karyotyping and microarrays for balanced and unbalanced chromosome rearrangements, respectively, and southern blot and repeat-primed PCR for the amplification and sizing of expanded alleles, impaired by limited resolution and sensitivity that have not been significantly improved by the advent of NGS. Nevertheless, more recently, with the increased accuracy provided by the latest product releases, LRS has been tested also for SNV detection, especially in genes with highly homologous pseudogenes and for haplotype reconstruction to assess the parental origin of alleles with de novo pathogenic variants. We provide a review of relevant recent scientific papers exploring LRS potential in the diagnosis of genetic diseases and its potential future applications in routine genetic testing. |
| first_indexed | 2024-03-07T14:30:43Z |
| format | Article |
| id | doaj.art-1824b60ae34e4ed3918ab0901427b3ca |
| institution | Directory Open Access Journal |
| issn | 1664-8021 |
| language | English |
| last_indexed | 2024-03-07T14:30:43Z |
| publishDate | 2024-03-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Genetics |
| spelling | doaj.art-1824b60ae34e4ed3918ab0901427b3ca2024-03-06T05:04:54ZengFrontiers Media S.A.Frontiers in Genetics1664-80212024-03-011510.3389/fgene.2024.13748601374860Long read sequencing on its way to the routine diagnostics of genetic diseasesGiulia Olivucci0Giulia Olivucci1Emanuela Iovino2Giovanni Innella3Giovanni Innella4Daniela Turchetti5Daniela Turchetti6Tommaso Pippucci7Pamela Magini8IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyDepartment of Surgical and Oncological Sciences, University of Palermo, Palermo, ItalyIRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyDepartment of Medical and Surgical Sciences (DIMEC), University of Bologna, Bologna, ItalyMedical Genetics Unit, IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyDepartment of Medical and Surgical Sciences (DIMEC), University of Bologna, Bologna, ItalyMedical Genetics Unit, IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyIRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyMedical Genetics Unit, IRCCS Azienda Ospedaliero-Universitaria di Bologna, Bologna, ItalyThe clinical application of technological progress in the identification of DNA alterations has always led to improvements of diagnostic yields in genetic medicine. At chromosome side, from cytogenetic techniques evaluating number and gross structural defects to genomic microarrays detecting cryptic copy number variants, and at molecular level, from Sanger method studying the nucleotide sequence of single genes to the high-throughput next-generation sequencing (NGS) technologies, resolution and sensitivity progressively increased expanding considerably the range of detectable DNA anomalies and alongside of Mendelian disorders with known genetic causes. However, particular genomic regions (i.e., repetitive and GC-rich sequences) are inefficiently analyzed by standard genetic tests, still relying on laborious, time-consuming and low-sensitive approaches (i.e., southern-blot for repeat expansion or long-PCR for genes with highly homologous pseudogenes), accounting for at least part of the patients with undiagnosed genetic disorders. Third generation sequencing, generating long reads with improved mappability, is more suitable for the detection of structural alterations and defects in hardly accessible genomic regions. Although recently implemented and not yet clinically available, long read sequencing (LRS) technologies have already shown their potential in genetic medicine research that might greatly impact on diagnostic yield and reporting times, through their translation to clinical settings. The main investigated LRS application concerns the identification of structural variants and repeat expansions, probably because techniques for their detection have not evolved as rapidly as those dedicated to single nucleotide variants (SNV) identification: gold standard analyses are karyotyping and microarrays for balanced and unbalanced chromosome rearrangements, respectively, and southern blot and repeat-primed PCR for the amplification and sizing of expanded alleles, impaired by limited resolution and sensitivity that have not been significantly improved by the advent of NGS. Nevertheless, more recently, with the increased accuracy provided by the latest product releases, LRS has been tested also for SNV detection, especially in genes with highly homologous pseudogenes and for haplotype reconstruction to assess the parental origin of alleles with de novo pathogenic variants. We provide a review of relevant recent scientific papers exploring LRS potential in the diagnosis of genetic diseases and its potential future applications in routine genetic testing.https://www.frontiersin.org/articles/10.3389/fgene.2024.1374860/fulllong read sequencingmolecular diagnosisgenetic diseasesstructural variantstandem repeatssingle nucleotide variants |
| spellingShingle | Giulia Olivucci Giulia Olivucci Emanuela Iovino Giovanni Innella Giovanni Innella Daniela Turchetti Daniela Turchetti Tommaso Pippucci Pamela Magini Long read sequencing on its way to the routine diagnostics of genetic diseases Frontiers in Genetics long read sequencing molecular diagnosis genetic diseases structural variants tandem repeats single nucleotide variants |
| title | Long read sequencing on its way to the routine diagnostics of genetic diseases |
| title_full | Long read sequencing on its way to the routine diagnostics of genetic diseases |
| title_fullStr | Long read sequencing on its way to the routine diagnostics of genetic diseases |
| title_full_unstemmed | Long read sequencing on its way to the routine diagnostics of genetic diseases |
| title_short | Long read sequencing on its way to the routine diagnostics of genetic diseases |
| title_sort | long read sequencing on its way to the routine diagnostics of genetic diseases |
| topic | long read sequencing molecular diagnosis genetic diseases structural variants tandem repeats single nucleotide variants |
| url | https://www.frontiersin.org/articles/10.3389/fgene.2024.1374860/full |
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