Aditya Singh1, Prateek Bhatia1. 1. Paediatric Haematology-Oncology Unit, Post Graduate Institute of Medical Education and Research, Chandigarh, India.
Abstract
INTRODUCTION: Currently, two second-generation sequencing platforms-Ion Torrent and Illumina are being widely used for clinical testing and reporting of human samples. However, third-generation long read platforms like single molecule (PacBio) or direct sequencing (Nanopore) are gaining widespread interest in clinical genomics. AIMS AND OBJECTIVES: The current study attempts to analyse and compare MinION sequencing data with Ion Torrent data, using a haematology ampliseq panel, to shed light on its current standing in reporting of clinical diagnostic samples. METHODOLOGY: A custom targeted Next-generation sequencing ampliseq panel comprising of 33 genes related to detection of inherited bone marrow failure syndrome cases was used to sequence five samples on both Ion Torrent and Oxford MinION platforms. The resulting data were analysed for output, quality and variant metrics across both platforms independently. RESULTS: Overall, MinION produced longer reads (range 108-7227 bp) than Ion Torrent (25-580 bp). Moreover, it generated more reads than Ion Torrent in high %GC content (P < .005) or repeat regions. But Ion Torrent had much lower error rate of 1.59% compared with MinION's 20.31%. Despite high error, MinION platform was able to identify and report the pathogenic variant in all samples. DISCUSSION AND CONCLUSION: The extremely long read lengths of MinION sequencers and better coverage in difficult to sequence regions give it an edge in generating contig free whole-genome sequences. However, the pore technology and chemistry of MinION needs further tuning to reduce error rate before it can be incorporated for clinical testing and reporting of human samples.
INTRODUCTION: Currently, two second-generation sequencing platforms-Ion Torrent and Illumina are being widely used for clinical testing and reporting of human samples. However, third-generation long read platforms like single molecule (PacBio) or direct sequencing (Nanopore) are gaining widespread interest in clinical genomics. AIMS AND OBJECTIVES: The current study attempts to analyse and compare MinION sequencing data with Ion Torrent data, using a haematology ampliseq panel, to shed light on its current standing in reporting of clinical diagnostic samples. METHODOLOGY: A custom targeted Next-generation sequencing ampliseq panel comprising of 33 genes related to detection of inherited bone marrow failure syndrome cases was used to sequence five samples on both Ion Torrent and Oxford MinION platforms. The resulting data were analysed for output, quality and variant metrics across both platforms independently. RESULTS: Overall, MinION produced longer reads (range 108-7227 bp) than Ion Torrent (25-580 bp). Moreover, it generated more reads than Ion Torrent in high %GC content (P < .005) or repeat regions. But Ion Torrent had much lower error rate of 1.59% compared with MinION's 20.31%. Despite high error, MinION platform was able to identify and report the pathogenic variant in all samples. DISCUSSION AND CONCLUSION: The extremely long read lengths of MinION sequencers and better coverage in difficult to sequence regions give it an edge in generating contig free whole-genome sequences. However, the pore technology and chemistry of MinION needs further tuning to reduce error rate before it can be incorporated for clinical testing and reporting of human samples.
Authors: Imogen A Wright; Kayla E Delaney; Mary Grace K Katusiime; Johannes C Botha; Susan Engelbrecht; Mary F Kearney; Gert U van Zyl Journal: Cells Date: 2021-09-28 Impact factor: 6.600