Nanopore vs. Illumina: Which DNA sequencing technology suits your project?
Read our Nanopore vs. Illumina analysis to compare their sequencing processes, understand their strengths and limitations, and determine the best technology.
January 1, 2025
Genetics has come a long way over the last three decades. The first attempt at human genome sequencing, which involved determining the exact sequence of DNA bases, took over a decade to complete. Today, however, advanced technologies can accomplish the same task in less than two days with even greater accuracy.
Two prominent methods currently in use are Illumina’s next-generation sequencing and nanopore sequencing by Oxford Nanopore Technologies. Both technologies are highly popular and respected but typically serve different use cases.
To determine the best option for your project needs, read our Nanopore vs. Illumina comparison to understand their unique strengths and weaknesses. We’ll explain how their sequencing processes differ and how they perform regarding throughput, accuracy, and cost-efficiency.
Illumina: Industry-standard, next-generation sequencing
Illumina is a leading company in the field of sequencing, with a revenue of $1.09 billion as of the second quarter of 2024. The company has played a key role in the widespread success of short-read, next-generation sequencing (NGS) after acquiring the company that developed the technology in the early 2000s.
NGS technology was revolutionary because it was faster and more efficient than its predecessor, Sanger sequencing. For details, read our Illumina vs. Sanger comparison.
Through its continuous efforts to increase throughput, Illumina has doubled its data output each year, reduced costs, and made DNA sequencing more accessible to the wider masses.
Illumina has built a robust infrastructure, workflow, and community, providing valuable tools and resources that many scientists swear by. The company sells a wide range of instruments, software, and preparation kits for different use cases, including common ones like:
Microarray-based genotyping that tests like 23andMe and AncestryDNA use
Whole-genome sequencing, the most advanced sequencing method available
As a continuous innovator, Illumina has recently expanded beyond its signature short-read technology and now offers machines that enable both short and long-read sequencing.
Source: Illumina via YouTube
Nanopore: New and unique approach to sequencing
Over the last decade, we’ve seen the rise of third-generation sequencing, which can read longer strands of DNA. Pacific Biosciences (PacBio) popularized this method, but Oxford Nanopore Technologies (ONT) released its own proprietary method soon after.
Oxford researchers developed the idea for this method, called nanopore sequencing, in the 1990s, but it wasn’t commercialized until the 2010s. Nanopore sequencing was unique because it relied on an entirely different approach than its predecessors.
Instead of fluorescent labeling, nanopore uses electrical current to detect the DNA sequence, enabling faster, real-time analysis. Nanopore can also read any length of fragments, from short to ultra-long.
Another standout feature of ONT is its pocket-sized MinION—the only portable sequencer available today. The company also sells various other high-throughput machines, prep kits, and software.
Source: Oxford Nanopore Technologies
Bonus read: Check out our detailed PacBio vs. Illumina sequencing breakdown.
Illumina vs. Nanopore sequencing: A closer look
Both Illumina and Nanopore can sequence DNA and RNA, offer high throughput, and are esteemed methods used by scientists everywhere. Still, they differ in almost every other way.
Here’s a brief overview of their basic characteristics before we examine their differences:
In the next sections, we’ll explain and compare Illumina and Nanopore sequencing in greater detail, focusing on these crucial factors:
Sequencing process
Result accuracy
Efficiency and convenience
Potential cost
Applications
1. Sequencing process
Illumina’s approach, called sequencing by synthesis (SBS), is a short-read method that analyzes small DNA fragments with high efficiency and accuracy.
The process goes as follows: Illumina first breaks down the DNA into fragments and amplifies them to create strands. Next, it repeats the process many times to form clusters identical to the original molecule. To identify the sequence of nucleotides, the DNA building blocks, Illumina labels them with a fluorescent dye before adding them to the amplified fragments one by one.
For long-read sequencing, SBS is accompanied by the secondary analysis of the DRAGEN (Dynamic Read Analysis for GENomics). With this approach, Illumina allows for a streamlined short and long-read workflow within one sequencer rather than requiring a separate machine or process.
Source: Illumina
Nanopore sequencing is a method that can analyze both short and ultra-long fragments of DNA, offering much greater long-read coverage than Illumina.
In this process, individual DNA molecules pass through nanopores, tiny cavities in the electro-resistant membrane of flow or sample cells. These nanopores can detect the electric current of the molecules that flow through them. Using basecalling algorithms, the machine can decode these disruptions to identify nucleotides and determine the DNA sequence in real time.
2. Result accuracy
Although not the most precise analysis available, Illumina sequencing is generally highly accurate. We typically evaluate the quality of sequencing data and the probability of error using the industry-standard Phred scores. Most Illumina bases score Q30 or above — meaning they’re at least 99.9% accurate.
Illumina typically entails a low error rate, though sources have reported occasional:
Amplification biases
Indel (insertion and deletion) errors
Issues with sequence motifs and GC (guanine and cytosine)-rich regions
Source: Illumina
Nanopore is generally accurate, but it’s more error-prone than Illumina sequencing. Its quality varies significantly, with sources reporting Phred scores ranging from around Q10 to Q25. Nanopore sequencing seems to struggle with the following:
Distinguishing homopolymeric regions
Systematic errors
Indel errors
That said, Nanopore’s accuracy has improved over the years and is likely to continue doing so. According to ONT, the latest Dorado basecalling model now allows it to achieve the Q26 benchmark, or 99.75% accuracy.
3. Efficiency
Illumina sequencing is known for being highly efficient and scalable. Its system NovaSeq X Plus can reach ultra-high throughput, delivering up to 16 Tb of output per dual run.
As it can analyze millions of DNA fragments and multiple samples at a time, Illumina is usually the preferred option for large and demanding projects. Illumina can now analyze the entire human genome in under 30 hours and is constantly working on reducing the time, effort, and cost of the prep work and overall workflow.
Nanopore sequencing doesn’t fall far behind, though. Like Illumina, it’s scalable and can read both short and long fragments. Nanopore sequencing can also sequence DNA in real time, which reduces the need for post-sequencing data processing and provide rapid insights.
Sources report that Nanopore can complete a whole-genome sequence within two hours and identify causative genetic variants in eight — making the service the fastest option available.
Source: Oxford Nanopore Technologies
4. Potential cost
The cost-efficiency of either method varies greatly and depends on various factors, largely the specific device and use case. Nanopore’s instruments offer flexible read length and throughput, so they can serve many purposes and minimize the upfront investment.
Some of Illumina’s instruments offer both short and long reads, but its ability to generate vast amounts of accurate data makes it among the most economical sequencing methods available. This applies to both the cost per run and the cost per gigabyte.
It’s also worth noting that Illumina is the reason genomic testing is accessible to a wide audience today. Numerous DNA companies, including MyHeritage, tellmeGen, and Sequencing.com, use Illumina’s technology to offer affordable genetic tests for ancestry and health.
Still, the most notable example is Nucleus — a platform offering all-in-one DNA health insights. With Illumina’s technology, Nucleus can provide whole-genome tests for only $399, while many other companies charge thousands. The test can deliver life-changing insights, including your risk for various common and rare diseases, carrier status, and genetic traits.
5. Applications
Illumina and Nanopore sequencing have unique strengths and are thus used for different types of projects. Due to its flexible throughput and portability, ONT has a broader range of applications, though Illumina’s precision is sometimes preferred.
Discover the optimal use cases of these sequencing methods below:
Illumina sequencing is ideal for…Nanopore sequencing is ideal for…Large-scale projects, such as genome-wide association studiesClinical diagnostics that require high accuracy and reliability, such as cancer genomicsGenome or DNA region assemblyDetection of structural variants and repetitive DNA areasRapid diagnosticsFieldwork and in situ analysis
As many labs have an established Illumina infrastructure and get extensive support from the company, they often find it challenging and unprofitable to switch to a completely different system like Nanopore.
Source: Oxford Nanopore Technologies
Nanopore vs. Illumina: Final thoughts
Considering Illumina and Nanopore sequencing are highly respected methods with distinct benefits, assessing which one is better is only possible by knowing the intent.
Illumina’s biggest strengths are its accuracy, extensive output, and proven infrastructure, while Nanopore stands out for its ultra-long reads, flexible throughput, and portability. To make your decision, you should consider your project requirements, budget, and existing infrastructure.
That said, it’s important to remember that Illumina is responsible for the widespread accessibility of genomic testing today. Its high-throughput sequencing method was groundbreaking, and today, it allows companies like Nucleus to provide at-home genetic tests at an affordable cost.
These crucial insights give more people agency over their well-being, guiding their choices toward a brighter future for themselves and their families.
Nucleus: One genetic test, numerous key insights
Nucleus is an all-in-one DNA platform that offers physician-ordered tests. Based on a cheek swab, Nucleus can analyze nearly all six billion data points of your genome, including rare genetic differences that other tests miss but can have a significant impact.
You can learn if your genes predispose you to over 800 conditions and diseases, including:
Physical conditions such as heart disease, rare cancers, and Alzheimer’s disease
Mental health conditions such as depression, ADHD, and schizophrenia
To further customize your reports, Nucleus will consider non-genetic factors, such as your BMI, as they can modify your gene expression. You can even find out how your genes influence traits like intelligence, muscle strength, and longevity.
Nucleus’ reports are easy to follow for everyone as they use simple language and illustrations. If you want expert advice regarding your results, you can request a session with a genetic counselor through Nucleus’ partner, SteadyMD.
A clinical-grade platform, Nucleus is fully regulated, works with certified U.S. labs, and uses the latest Illumina technology to analyze your DNA with 99.9% accuracy. The company is also HIPAA-compliant, ensuring the safety and privacy of your sensitive data.
Own your health with Nucleus Premium
To have your whole DNA analyzed, purchase the Nucleus Premium test kit first. Here’s how you can do that:
Provide your personal information
Place the order
The kit costs $399 and is HSA and FSA-eligible. Once you swab your cheeks and mail in the samples, you’ll get your health and trait reports online in six to eight weeks. Nucleus has an ever-expanding database and will send updated reports in light of advancements as well as your lifestyle changes.
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Featured image source: alanajordan