Autor(es):
Silva, Catarina ; Ferrão, José ; Marques, Bárbara ; Pedro, Sónia ; Correia, Hildeberto ; Valente, Ana ; Rodrigues, Antonio Sebastião ; L, Vieira
Data: 2025
Identificador Persistente: http://hdl.handle.net/10362/188427
Origem: Repositório Institucional da UNL
Assunto(s): Copy number variation; Hybrid-SNP microarray; Nanopore Sequencing; Structural variation; Third-generation sequencing; Variant calling; Whole genome sequencing; Biochemistry; Molecular Medicine; Molecular Biology; Genetics; Genetics(clinical); Biochemistry, medical
Descrição
Funding Information: This work is a result of the GenomePT project (POCI-01\u20130145-FEDER-022184), supported by COMPETE 2020\u2014Operational Programme for Competitiveness and Internationalization (POCI), Lisboa Portugal Regional Operational Programme (Lisboa2020), Algarve Portugal Regional Operational Programme (CRESC Algarve2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF), and by Funda\u00E7\u00E3o para a Ci\u00EAncia e a Tecnologia (FCT). This work was also supported by Fundos FEDER through the Programa Operacional Factores de Competitividade \u2013 COMPETE and by Fundos Nacionais through the Funda\u00E7\u00E3o para a Ci\u00EAncia e a Tecnologia within the scope of the project UID/BIM/00009/2019 (Centre for Toxicogenomics and Human Health -ToxOmics). Publisher Copyright: © The Author(s) 2025.
Background: Nanopore sequencing is a technology that holds great promise for identifying all types of human genome variations, particularly structural variations. In this work, we used nanopore sequencing technology to sequence 2 human cell lines at low depth of coverage to call copy number variations (CNV), and compared the results variant by variant with chromosomal microarray (CMA) results. Results: We analysed sequencing data using CuteSV and Sniffles2 variant callers, compared breakpoints based on hybrid-SNP microarray, nanopore sequencing and Sanger sequencing, and analysed CNV coverage. From a total of 48 high confidence variants (truth set), variant calling detected 79% of the truth set variants, increasing to 86% for interstitial CNV. Simultaneous use of the 2 callers slightly increased variant calling. Both callers performed better when calling CNV losses than gains. Variant sizes from CMA and nanopore sequencing showed an excellent correlation, with breakpoints determined by nanopore sequencing differing by only 20 base pairs on average from Sanger sequencing. Nanopore sequencing also revealed that four variants concealed genomic inversions undetectable by CMA. In the 10 CNV not called in nanopore sequencing, 8 showed coverage evidence of genomic loss or gain, highlighting the need to improve SV calling algorithms performance. Conclusions: Nanopore sequencing offers advantages over CMA for structural variant detection, including the identification of multiple variant types and their breakpoints with increased precision. However, further improvements in variant calling algorithms are still needed for nanopore sequencing to become a highly robust and standardized approach for a comprehensive analysis of genomic structural variation.
