3-hour genome sequencing and targeted analysis to rapidly assess genetic risk
Purpose: Rapid genetic testing in the critical care setting may guide diagnostic evaluation, direct therapies, and help families and care providers make informed decisions about goals of care. We tested whether a simplified DNA extraction and library preparation process would enable us to perform ul...
| Main Authors: | , , , , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2024-01-01
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| Series: | Genetics in Medicine Open |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2949774424009798 |
| _version_ | 1797220203362254848 |
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| author | Miranda P.G. Zalusky Jonas A. Gustafson Stephanie C. Bohaczuk Ben Mallory Paxton Reed Tara Wenger Erika Beckman Irene J. Chang Cate R. Paschal Jillian G. Buchan Christina M. Lockwood Mihai Puia-Dumitrescu Daniel R. Garalde Joseph Guillory Androo J. Markham Michael J. Bamshad Evan E. Eichler Andrew B. Stergachis Danny E. Miller |
| author_facet | Miranda P.G. Zalusky Jonas A. Gustafson Stephanie C. Bohaczuk Ben Mallory Paxton Reed Tara Wenger Erika Beckman Irene J. Chang Cate R. Paschal Jillian G. Buchan Christina M. Lockwood Mihai Puia-Dumitrescu Daniel R. Garalde Joseph Guillory Androo J. Markham Michael J. Bamshad Evan E. Eichler Andrew B. Stergachis Danny E. Miller |
| author_sort | Miranda P.G. Zalusky |
| collection | DOAJ |
| description | Purpose: Rapid genetic testing in the critical care setting may guide diagnostic evaluation, direct therapies, and help families and care providers make informed decisions about goals of care. We tested whether a simplified DNA extraction and library preparation process would enable us to perform ultrarapid assessment of genetic risk for a Mendelian condition, based on information from an affected sibling, using long-read genome sequencing and targeted analysis. Methods: Following extraction of DNA from cord blood and rapid library preparation, genome sequencing was performed on an Oxford Nanopore PromethION. FASTQ files were generated from original sequencing data in near real-time and aligned to a reference genome. Variant calling and analysis were performed at timed intervals. Results: We optimized the DNA extraction and library preparation methods to create sufficient library for sequencing from 500 μL of blood. Real-time, targeted analysis was performed to determine that the newborn was neither affected nor a heterozygote for variants underlying a Mendelian condition. Phasing of the target region and prior knowledge of the affected haplotypes supported our interpretation despite a low level of coverage at 3 hours of life. Conclusion: This proof-of-concept experiment demonstrates how prior knowledge of haplotype structure or familial variants can be used to rapidly evaluate an individual at risk for a genetic disease. Although ultrarapid sequencing remains both complex and cost prohibitive, our method is more easily automated than prior approaches and uses smaller volumes of blood and thus may be more easily adopted for future studies of ultrarapid genome sequencing in the clinical setting. |
| first_indexed | 2024-04-24T12:45:48Z |
| format | Article |
| id | doaj.art-6d44b504cf61496087437a8f47863416 |
| institution | Directory Open Access Journal |
| issn | 2949-7744 |
| language | English |
| last_indexed | 2024-04-24T12:45:48Z |
| publishDate | 2024-01-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Genetics in Medicine Open |
| spelling | doaj.art-6d44b504cf61496087437a8f478634162024-04-07T04:37:06ZengElsevierGenetics in Medicine Open2949-77442024-01-0121018333-hour genome sequencing and targeted analysis to rapidly assess genetic riskMiranda P.G. Zalusky0Jonas A. Gustafson1Stephanie C. Bohaczuk2Ben Mallory3Paxton Reed4Tara Wenger5Erika Beckman6Irene J. Chang7Cate R. Paschal8Jillian G. Buchan9Christina M. Lockwood10Mihai Puia-Dumitrescu11Daniel R. Garalde12Joseph Guillory13Androo J. Markham14Michael J. Bamshad15Evan E. Eichler16Andrew B. Stergachis17Danny E. Miller18Division of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WA; Molecular and Cellular Biology Program, University of Washington School of Medicine, Seattle, WADivision of Medical Genetics, Department of Medicine, University of Washington, Seattle, WADivision of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA; Department of Genome Sciences, University of Washington School of Medicine, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WADepartment of Laboratory Medicine and Pathology, University of Washington, Seattle, WA; Department of Laboratories, Seattle Children’s Hospital, Seattle, WADepartment of Laboratory Medicine and Pathology, University of Washington, Seattle, WADepartment of Genome Sciences, University of Washington School of Medicine, Seattle, WA; Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA; Brotman Baty Institute for Precision Medicine, University of Washington, Seattle, WADivision of Neonatology, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WAOxford Nanopore Technologies, Oxford, United KingdomOxford Nanopore Technologies, Oxford, United KingdomOxford Nanopore Technologies, Oxford, United KingdomDivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WA; Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA; Brotman Baty Institute for Precision Medicine, University of Washington, Seattle, WADepartment of Genome Sciences, University of Washington School of Medicine, Seattle, WA; Brotman Baty Institute for Precision Medicine, University of Washington, Seattle, WA; Howard Hughes Medical Institute, University of Washington, Seattle, WADivision of Medical Genetics, Department of Medicine, University of Washington, Seattle, WA; Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA; Brotman Baty Institute for Precision Medicine, University of Washington, Seattle, WADivision of Genetic Medicine, Department of Pediatrics, University of Washington and Seattle Children’s Hospital, Seattle, WA; Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA; Brotman Baty Institute for Precision Medicine, University of Washington, Seattle, WA; Correspondence and requests for materials should be addressed to Danny E. Miller, 1959 NE Pacific St, HSB H474A, Seattle, WA 98195.Purpose: Rapid genetic testing in the critical care setting may guide diagnostic evaluation, direct therapies, and help families and care providers make informed decisions about goals of care. We tested whether a simplified DNA extraction and library preparation process would enable us to perform ultrarapid assessment of genetic risk for a Mendelian condition, based on information from an affected sibling, using long-read genome sequencing and targeted analysis. Methods: Following extraction of DNA from cord blood and rapid library preparation, genome sequencing was performed on an Oxford Nanopore PromethION. FASTQ files were generated from original sequencing data in near real-time and aligned to a reference genome. Variant calling and analysis were performed at timed intervals. Results: We optimized the DNA extraction and library preparation methods to create sufficient library for sequencing from 500 μL of blood. Real-time, targeted analysis was performed to determine that the newborn was neither affected nor a heterozygote for variants underlying a Mendelian condition. Phasing of the target region and prior knowledge of the affected haplotypes supported our interpretation despite a low level of coverage at 3 hours of life. Conclusion: This proof-of-concept experiment demonstrates how prior knowledge of haplotype structure or familial variants can be used to rapidly evaluate an individual at risk for a genetic disease. Although ultrarapid sequencing remains both complex and cost prohibitive, our method is more easily automated than prior approaches and uses smaller volumes of blood and thus may be more easily adopted for future studies of ultrarapid genome sequencing in the clinical setting.http://www.sciencedirect.com/science/article/pii/S2949774424009798Genetic testingGenomicsLong-read sequencingNanoporeUltrarapid sequencing |
| spellingShingle | Miranda P.G. Zalusky Jonas A. Gustafson Stephanie C. Bohaczuk Ben Mallory Paxton Reed Tara Wenger Erika Beckman Irene J. Chang Cate R. Paschal Jillian G. Buchan Christina M. Lockwood Mihai Puia-Dumitrescu Daniel R. Garalde Joseph Guillory Androo J. Markham Michael J. Bamshad Evan E. Eichler Andrew B. Stergachis Danny E. Miller 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk Genetics in Medicine Open Genetic testing Genomics Long-read sequencing Nanopore Ultrarapid sequencing |
| title | 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| title_full | 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| title_fullStr | 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| title_full_unstemmed | 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| title_short | 3-hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| title_sort | 3 hour genome sequencing and targeted analysis to rapidly assess genetic risk |
| topic | Genetic testing Genomics Long-read sequencing Nanopore Ultrarapid sequencing |
| url | http://www.sciencedirect.com/science/article/pii/S2949774424009798 |
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