Interest of exome sequencing trio-like strategy based on pooled parental DNA for diagnosis and translational research in rare diseases.
Genetic Markers
Genetic Predisposition to Disease
Genetic Testing
Genome-Wide Association Study
Humans
Rare Diseases
/ diagnosis
Reproducibility of Results
Research Design
Sensitivity and Specificity
Sequence Analysis, DNA
Translational Research, Biomedical
/ methods
Exome Sequencing
/ methods
Workflow
cost effectiveness
exome sequencing
rare diseases
trio-like strategy; parental-pool strategy
Journal
Molecular genetics & genomic medicine
ISSN: 2324-9269
Titre abrégé: Mol Genet Genomic Med
Pays: United States
ID NLM: 101603758
Informations de publication
Date de publication:
12 2021
12 2021
Historique:
revised:
22
09
2021
received:
31
05
2021
accepted:
01
10
2021
pubmed:
31
10
2021
medline:
24
3
2022
entrez:
30
10
2021
Statut:
ppublish
Résumé
Exome sequencing (ES) has become the most powerful and cost-effective molecular tool for deciphering rare diseases with a diagnostic yield approaching 30%-40% in solo-ES and 50% in trio-ES. We applied an innovative parental DNA pooling method to reduce the parental sequencing cost while maintaining the diagnostic yield of trio-ES. We pooled six (Agilent-CRE-v2-100X) or five parental DNA (TWIST-HCE-70X) aiming to detect allelic balance around 8-10% for heterozygous status. The strategies were applied as second-tier (74 individuals after negative solo-ES) and first-tier approaches (324 individuals without previous ES). The allelic balance of parental-pool variants was around 8.97%. Sanger sequencing uncovered false positives in 1.5% of sporadic variants. In the second-tier approach, we evaluated than two thirds of the Sanger validations performed after solo-ES (41/59-69%) would have been saved if the parental-pool segregations had been available from the start. The parental-pool strategy identified a causative diagnosis in 18/74 individuals (24%) in the second-tier and in 116/324 individuals (36%) in the first-tier approaches, including 19 genes newly associated with human disorders. Parental-pooling is an efficient alternative to trio-ES. It provides rapid segregation and extension to translational research while reducing the cost of parental and Sanger sequencing.
Sections du résumé
BACKGROUND
Exome sequencing (ES) has become the most powerful and cost-effective molecular tool for deciphering rare diseases with a diagnostic yield approaching 30%-40% in solo-ES and 50% in trio-ES. We applied an innovative parental DNA pooling method to reduce the parental sequencing cost while maintaining the diagnostic yield of trio-ES.
METHODS
We pooled six (Agilent-CRE-v2-100X) or five parental DNA (TWIST-HCE-70X) aiming to detect allelic balance around 8-10% for heterozygous status. The strategies were applied as second-tier (74 individuals after negative solo-ES) and first-tier approaches (324 individuals without previous ES).
RESULTS
The allelic balance of parental-pool variants was around 8.97%. Sanger sequencing uncovered false positives in 1.5% of sporadic variants. In the second-tier approach, we evaluated than two thirds of the Sanger validations performed after solo-ES (41/59-69%) would have been saved if the parental-pool segregations had been available from the start. The parental-pool strategy identified a causative diagnosis in 18/74 individuals (24%) in the second-tier and in 116/324 individuals (36%) in the first-tier approaches, including 19 genes newly associated with human disorders.
CONCLUSIONS
Parental-pooling is an efficient alternative to trio-ES. It provides rapid segregation and extension to translational research while reducing the cost of parental and Sanger sequencing.
Identifiants
pubmed: 34716697
doi: 10.1002/mgg3.1836
pmc: PMC8683640
doi:
Substances chimiques
Genetic Markers
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e1836Informations de copyright
© 2021 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals LLC.
Références
Am J Hum Genet. 2012 Oct 5;91(4):597-607
pubmed: 23040492
Nat Genet. 2010 Dec;42(12):1109-12
pubmed: 21076407
Clin Transl Sci. 2018 Jan;11(1):11-20
pubmed: 28796411
Am J Hum Genet. 2017 May 4;100(5):695-705
pubmed: 28475856
Mol Genet Genomic Med. 2021 Dec;9(12):e1836
pubmed: 34716697
Nat Genet. 2010 Jan;42(1):30-5
pubmed: 19915526
Nature. 2009 Sep 10;461(7261):272-6
pubmed: 19684571
Nat Genet. 2010 Oct;42(10):851-8
pubmed: 20818383
Sci Rep. 2016 Sep 27;6:33735
pubmed: 27670852
Eur J Hum Genet. 2019 Sep;27(9):1398-1405
pubmed: 30979967
Nat Genet. 2017 Jan;49(1):36-45
pubmed: 27841880
Am J Hum Genet. 2019 Aug 1;105(2):403-412
pubmed: 31303265
Mutat Res. 2018 May;809:24-31
pubmed: 29677560
Am J Hum Genet. 2017 Nov 2;101(5):664-685
pubmed: 29100083
J Mol Diagn. 2019 Jan;21(1):38-48
pubmed: 30577886
Epilepsia. 2019 Jan;60(1):155-164
pubmed: 30525188
Gene. 2019 Jun 5;700:168-175
pubmed: 30904718
Genet Med. 2018 Jun;20(6):645-654
pubmed: 29095811
Genet Med. 2015 Oct;17(10):774-81
pubmed: 25590979
BMC Med. 2017 Dec 6;15(1):213
pubmed: 29207974
Bioinformatics. 2010 Jun 15;26(12):i318-24
pubmed: 20529923
Hum Mol Genet. 2003 Dec 1;12(23):3173-80
pubmed: 14532329
N Engl J Med. 2016 Jun 9;374(23):2246-55
pubmed: 27276562
Eur J Hum Genet. 2019 Oct;27(10):1519-1531
pubmed: 31231135
Curr Protoc Hum Genet. 2014 Apr 24;81:7.23.1-21
pubmed: 24763994
Hum Mutat. 2015 Oct;36(10):915-21
pubmed: 26295439
Am J Hum Genet. 2018 Feb 1;102(2):266-277
pubmed: 29395073
Nature. 2017 Feb 23;542(7642):433-438
pubmed: 28135719
Annu Rev Genomics Hum Genet. 2020 Aug 31;21:351-372
pubmed: 32283948
Am J Hum Genet. 2019 Apr 4;104(4):721-730
pubmed: 30929742
Genet Med. 2013 Sep;15(9):733-47
pubmed: 23887774
JAMA Pediatr. 2017 Sep 1;171(9):855-862
pubmed: 28759686
Genet Med. 2018 Dec;20(12):1617-1626
pubmed: 29789557
NPJ Genom Med. 2018 Jul 9;3:16
pubmed: 30002876
Genome Med. 2017 Mar 21;9(1):26
pubmed: 28327206
Proc Natl Acad Sci U S A. 2015 Apr 28;112(17):5473-8
pubmed: 25827230
Eur J Hum Genet. 2017 Dec;25(12):1364-1376
pubmed: 29158550
Lancet. 2012 Nov 10;380(9854):1674-82
pubmed: 23020937
J Adv Res. 2020 Jan 30;23:113-121
pubmed: 32099673
Hum Genet. 2020 Nov;139(11):1381-1390
pubmed: 32399599
Eur J Hum Genet. 2018 Jan;26(1):54-63
pubmed: 29209020
Genet Med. 2018 Dec;20(12):1564-1574
pubmed: 29595814
Genet Med. 2016 Nov;18(11):1090-1096
pubmed: 26938784