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Whole-genome long-read sequencing downsampling and its effect on variant-calling precision and recall
William T Harvey1, Peter Ebert2,3,4, Jana Ebler2,4
1Department of Genome Sciences, University of Washington School of Medicine, Seattle, Washington 98195-5065, USA.
Genome Research
|January 8, 2024
Summary
Long-read sequencing (LRS) offers improved genome assembly and variant detection. Both Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PacBio) HiFi platforms show good performance, with PacBio HiFi excelling in assembly-based variant calling.
Area of Science:
- Genomics
- Bioinformatics
- Molecular Biology
Background:
- Long-read sequencing (LRS) advances whole-genome sequencing, enabling phased de novo genome assembly and access to challenging genomic regions.
- LRS facilitates the discovery of complex structural variants (SVs) linked to diseases, surpassing short-read sequencing limitations.
- Despite progress, LRS faces challenges in cost, scalability, and platform-specific read accuracy, necessitating careful experimental design.
Purpose of the Study:
- To compare the genetic variant-calling precision and recall of Oxford Nanopore Technologies (ONT) and Pacific Biosciences (PacBio) HiFi platforms.
- To evaluate the impact of varying sequence coverage on LRS variant detection sensitivity and accuracy.
- To assess the benefits of genome assembly for improving SV and indel variant calling using LRS data.
Main Methods:
- Comparative analysis of variant-calling performance between ONT and PacBio HiFi platforms.
- Systematic variation of sequence coverage levels to determine optimal thresholds for LRS applications.
- Evaluation of F1 scores for read-based and assembly-based variant call sets to measure precision and recall.
Main Results:
- LRS sensitivity plateaus around 12-fold coverage, with most variants accurately called (F1 score > 0.5) for read-based applications.
- Both ONT and PacBio HiFi platforms demonstrate robust performance in structural variant detection.
- Genome assembly enhances variant-calling precision and recall for SVs and indels, with PacBio HiFi showing superior performance over ONT in assembly-based variant calls.
Conclusions:
- Cost-effective experimental strategies for LRS can be designed without compromising the discovery of novel biological insights.
- PacBio HiFi demonstrates an advantage over ONT in genome assembly quality and subsequent variant calling.
- Continued evolution of LRS technologies promises further improvements in genomic analyses.
Related Concept Videos
01:20Downsampling
When considering a sampled sequence with zero values between sampling instants, one can replace it by taking every N-th value of the sequence. At these integer multiples of N, the original and sampled sequences coincide. This process, known as decimation, involves extracting every N-th sample from a sequence, thereby creating a more efficient sequence.
The Fourier transform of the decimated sequence reveals a combination of scaled and shifted versions of the original spectrum. This...
The Fourier transform of the decimated sequence reveals a combination of scaled and shifted versions of the original spectrum. This...
01:22Upsampling
Managing signal sampling rates is essential in digital signal processing to maintain signal integrity. A decimated signal, characterized by a reduced frequency range due to its lower sampling rate, can be upsampled by inserting zeros between each sample. This upsampling process expands the original spectrum and introduces repeated spectral replicas at intervals dictated by the new Nyquist frequency. To refine this zero-inserted sequence, it is passed through a lowpass filter with a cutoff...