Benchmarking of Nanopore R10.4 and R9.4.1 flow cells in single-cell whole-genome amplification and whole-genome shotgun sequencing

Research output: Journal Publications and Reviews (RGC: 21, 22, 62)21_Publication in refereed journalpeer-review

8 Scopus Citations
View graph of relations

Detail(s)

Original languageEnglish
Pages (from-to)2352-2364
Number of pages13
Journal / PublicationComputational and Structural Biotechnology Journal
Volume21
Online published24 Mar 2023
Publication statusPublished - 2023

Link(s)

Abstract

Third-generation sequencing can be used in human cancer genomics and epigenomic research. Oxford Nanopore Technologies (ONT) recently released R10.4 flow cell, which claimed an improved read accuracy compared to R9.4.1 flow cell. To evaluate the benefits and defects of R10.4 flow cell for cancer cell profiling on MinION devices, we used the human non-small-cell lung-carcinoma cell line HCC78 to construct libraries for both single-cell whole-genome amplification (scWGA) and whole-genome shotgun sequencing. The R10.4 and R9.4.1 reads were benchmarked in terms of read accuracy, variant detection, modification calling, genome recovery rate and compared with the next generation sequencing (NGS) reads. The results highlighted that the R10.4 outperforms R9.4.1 reads, achieving a higher modal read accuracy of over 99.1%, superior variation detection, lower false-discovery rate (FDR) in methylation calling, and comparable genome recovery rate. To achieve high yields scWGA sequencing in the ONT platform as NGS, we recommended multiple displacement amplification with a modified T7 endonuclease Ⅰ cutting procedure as a promising method. In addition, we provided a possible solution to filter the likely false positive sites among the whole genome region with R10.4 by using scWGA sequencing result as a negative control. Our study is the first benchmark of whole genome single-cell sequencing using ONT R10.4 and R9.4.1 MinION flow cells by clarifying the capacity of genomic and epigenomic profiling within a single flow cell. A promising method for scWGA sequencing together with the methylation calling results can benefit researchers who work on cancer cell genomic and epigenomic profiling using third-generation sequencing.

© 2023 The Authors. Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.

Research Area(s)

  • Long read, Nanopore DNA sequencing, Methylation, Single-cell whole genome amplification sequencing, Whole genome shotgun sequencing

Bibliographic Note

© 2023 The Authors.

Citation Format(s)

Benchmarking of Nanopore R10.4 and R9.4.1 flow cells in single-cell whole-genome amplification and whole-genome shotgun sequencing. / Ni, Ying; Liu, Xudong; Simeneh, Zemenu Mengistie et al.
In: Computational and Structural Biotechnology Journal, Vol. 21, 2023, p. 2352-2364.

Research output: Journal Publications and Reviews (RGC: 21, 22, 62)21_Publication in refereed journalpeer-review

Download Statistics

No data available