June 25, 2024

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Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours

Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours


Stanford/Oxford Nanopore/Google/NVIDIA Collaboration to Develop Ultra-Fast Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours.

Whole genome sequencing (WGS) allows scientists to see a person’s complete DNA sequence makeup, which contains information on everything from eye color to genetic diseases.

Therefore, genome sequencing is critical for diagnosing diseases in patients that involve DNA—once doctors know a specific genetic mutation, they can plan treatment accordingly.


It often takes weeks to sequence a patient’s whole genome and analyze the results, which is considered rapid by most doctors. But for some acutely ill patients, this time is obviously still too long. So, can it be further reduced to days or even hours?


On March 28, 2022, Stanford University, in conjunction with the University of California, Santa Cruz Genome Institute, Google, Oxford Nanopore, and NVIDIA, published a paper entitled: Accelerated identification of disease-causing variants with ultra-rapid nanopore in the journal Nature Biotechnology Research papers on genome sequencing .


In this study, the research team developed an ultra-rapid nanopore whole- genome sequencing method (ultra-rapid nanopore WGS) that combines an optimized sample preparation protocol to distribute sequencing to 48 sequencing flow cells. ) synchronous work, near real-time base detection and alignment, to speed up the detection of mutation sites.


The research team applied the method to two clinical cases, and it took less than 8 hours from the preparation of the sequencing sample to the detection of the disease-causing gene mutation .

This demonstrates that this ultrafast nanopore whole-genome sequencing approach provides accurate detection of disease-causing genetic mutations, accelerating diagnostic clinical genome sequencing compared to previous methods.


Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours


Whole genome sequencing (WGS) has important advantages in clinical medical diagnosis, especially in intensive care settings, but the process of sequencing and post-sequencing data analysis is often slow, which limits symptomatic treatment for some acute patients.


Whole-genome sequencing usually uses next-generation sequencing (NGS) technology, which cuts the genome into small fragments, reads the DNA sequence of each fragment, and finally stitches the entire genome together using the standard human genome as a reference.

But the sequencing and data analysis of this method is time-consuming, doesn’t always capture the full extent of our genome, and the information it provides sometimes omits genetic variants that point to a diagnosis.


In recent years, nanopore sequencing technology has developed rapidly and has become a high-throughput, high-fidelity sequencing platform.

The PromethION platform developed by Oxford Nanopore can accommodate 48 sequencing flow cells (flow cells) , each of which can complete sequencing independently.


The research team speculates that the 48 flow cells are sequenced simultaneously, capable of sequencing a single sample to clinical-quality depth within 2 hours. In addition, the research team believes that the alignment and mutation site discovery after sequencing can be completed within hours.


But there are some obvious technical challenges ahead to get there. First, the traditional sequencing sample preparation scheme does not consider the rapid generation of sequencing libraries that meet the needs of clinical sequencing from limited blood samples; secondly, 48 sequencing flow cells are synchronously sequenced, which can achieve near real-time sequencing data output, but data generation The speed far exceeds the speed of base alignment; third, the performance of sequencing small mutations using nanopores has not been characterized in clinical samples; finally, traditional mutation filtering and prioritization methods lead to about 100 mutation candidates loci, which require manual further screening and confirmation, limiting efficiency.


In this study, the research team teamed up with the UC Santa Cruz Genome Institute, Google, Oxford Nanopore, and Nvidia to address these challenges.

They developed an ultra-rapid nanopore whole- genome sequencing method (ultra-rapid nanopore WGS) , which optimizes the method of gene sequencing sample preparation and uploads the data to the Google Cloud Computing Platform , with NVIDIA providing high-performance computing to perform near real-time Base calling and alignment, greatly accelerating the discovery of single nucleotide polymorphisms (SNPs) , insertion and deletion mutations (indels), and structural variants (SVs) , and using the Genome In A Bottle sample as a reference genome for comparison pair and characterize.


Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours


Finally, the research team used the method to sequence a diagnosis in a 57-year-old critically ill man and a 14-month-old infant less than 8 hours after blood draw (7 hours 18 minutes and 7 hours 48 minutes, respectively) Candidate gene mutations were detected. The fastest time reported to date (from sample preparation to mutation identification) is about 50% faster.


Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours



Overall, this study developed an ultra-rapid nanopore whole-genome sequencing method (ultra-rapid nanopore WGS) capable of deep sequencing of the human whole genome within 2 hours, combined with real-time base calling, alignment, Candidate disease-causing mutations were identified within 8 hours.


It is worth mentioning that the team published an article titled: Ultrarapid Nanopore Genome Sequencing in a Critical Care Setting in the New England Journal of Medicine (NEJM) on January 12, 2022 .


A research team has developed a new ultra-fast nanopore genome sequencing method for diagnosing rare genetic diseases in an average of 8 hours, unprecedented in standard clinical care.

Rapid diagnosis means patients spend less time in intensive care units, require fewer tests, recover faster and spend less on care. More importantly, this faster sequencing technology did not sacrifice accuracy, and the sequencing results were still reliable.


Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours


All in all, this ultra-fast nanopore whole-genome sequencing technology can diagnose a patient’s genetic cause in about eight hours and is a “game-changer” for genetic mutation diagnosis.


Professor Euan Ashley said: “Traditional standard tests screen patients’ blood for disease-related markers, but they scan only a few well-documented genes. This new technology can scan a patient’s entire genome for Any and all genetic variants, even if the gene was discovered the day before.”


After the publication of this study, more scientists began to pour into this field. Perhaps in the near future, genetic diagnosis will be like nucleic acid testing, and the results can be obtained in just a few hours, which is convenient for patients to get the fastest results. the most appropriate treatment.











Nanopore Sequencing: Discovering Disease-Causing Gene Mutations in 7 Hours

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