PacBio vs. Illumina: Which DNA sequencing technology will work for you?

With genomics research evolving rapidly over the last decade, we now have a couple of new DNA sequencing technologies to choose from — and they’re more efficient and affordable than ever. For many researchers and DNA enthusiasts, choosing the right technology is essential to ensuring the desired reporting depth and cost-to-value ratio. 

PacBio and Illumina are two leaders in the field of sequencing technology, and each has its own strengths and weaknesses, as well as niche use cases across DNA projects.

In this PacBio vs. Illumina analysis, we’ll give you a head-to-head comparison of the technologies offered by these biotechnology companies. You’ll learn about their sequencing methods, optimal use cases, and potential costs.

Bonus: We’ll also introduce you to Nucleus, a DNA platform that allows you to leverage the latest sequencing technology to unlock comprehensive health insights.

Illumina: Next-generation sequencing

As a company, Illumina has played a significant role in the massive success of next-generation sequencing (NGS) technology. NGS was developed in the early 2000s by a company that would later become part of Illumina, and has proved to be more efficient than its predecessor, Sanger sequencing. You can explore our Illumina vs. Sanger comparison here.

Over the years, Illumina’s short-read sequencing has become the most widely used method for DNA analysis. It has evolved quickly, with its output capacity increasing more than double each year. As a result, Illumina has managed to reduce the costs and turnaround time, making sequencing and genetic health and ancestry insights more widely accessible.

Today, the company sells a wide range of instruments and kits to support various DNA analyses, such as:

• Expression profiling

• RNA sequencing

• Microarray-based genotyping

• Whole-genome sequencing

Although Illumina specializes in short-read sequencing, it also launched a groundbreaking product that offers both short and long-read sequencing in 2023. 

Source: Illumina

Bonus read: Learn about popular DNA platforms that rely on Illumina sequencing technology:

• 23andMe review

• MyHeritage review

• Sequencing.com review

• tellmeGen review

PacBio: Third-generation sequencing 

About a decade after Illumina’s breakthrough, biotech company Pacific Biosciences (PacBio) brought in a new wave of long-read sequencing called third-generation sequencing. PacBio’s technology enabled more detailed analysis, higher throughput, and lower costs for long-read sequencing, much like Illumina did in the short-read sphere.

PacBio offers both long and short reads, though it’s best known for and specializes in long-read sequencing. Its long-read technology can sequence large and complex genomic sections easily — often more difficult to map using short-read technology.

PacBio also sells instruments and consumables, spanning use cases such as whole-genome, RNA, and targeted sequencing, as well as epigenetics. It has teamed up with DNA platforms like Invitae to make its testing technology available to more customers.

Source: PacBio

Illumina vs. PacBio: A closer look

Besides being able to thoroughly analyze your DNA, Illumina and PacBio sequencing have little in common, and they often serve different use cases altogether. Check out a brief overview of their characteristics before we get into the details:

| Aspect                       | Illumina                               | PacBio                            |
| ---------------------------- | -------------------------------------- | --------------------------------- |
| Founded in                   | 1998                                   | 2004                              |
| Main focus                   | Short-read sequencing                  | Long-read sequencing              |
| Short-read sequencing length  | 50–500 base pairs                      | 100–200 base pairs                |
| Short-read processing         | Sequencing by synthesis (SBS)          | Sequencing by binding (SBB)       |
| Short-read accuracy          | 99.90%                                 | 99.99%                            |
| Long-read sequencing length  | 5–10+ kilobase                        | 1–25 kilobase                     |
| Long-read processing         | Sequencing by synthesis (SBS) + DRAGEN | Single molecule, real-time (SMRT)

In the following sections, we’ll break down the key differentiating factors between Illumina and PacBio that can influence your choice:

1. Technology

2. Accuracy

3. Efficiency

4. Potential cost

5. Applications

1. Technology 

PacBio and Illumina rely on different methods to analyze DNA bases. In Illumina’s trademark short-read process, called sequencing by synthesis (SBS), the DNA is broken down into smaller fragments, which are then copied to form strands. 

Next, nucleotides are added one by one and are labeled with a fluorescent marker, which allows the machine to identify them and determine their order, i.e., the DNA sequence. Illumina’s method was groundbreaking as it enabled the sequencing of millions of fragments in parallel, while its predecessor, Sanger, could only read one at a time.

To enable both short and long reads from a single platform and improve accuracy, Illumina combines SBS with DRAGEN (short for Dynamic Read Analysis for GENomics) for secondary analysis.

Source: Illumina

On the other hand, PacBio has developed and uses the single molecule, real-time sequencing (SMRT) — a long-read sequencing method. In this method, the DNA is broken down into long fragments, which are then sequenced individually in real time without the need for amplification. Recently, it has launched High Fidelity (HiFi) reading, a more precise SMRT analysis.

For short reads, PacBio uses sequencing by binding (SBB), which also detects nucleotides continuously without amplification.

2. Accuracy 

To assess the accuracy of Illumina and PacBio sequencing, we’ve considered their Phred scores. These are standard measures used to assess the quality of identification of DNA sequencing data. Higher scores indicate a lower probability of reading error, whereas lower scores signal that a significant portion of the reading is unusable and may lead to false conclusions.

Both Illumina and PacBio sequencing methods are considered highly accurate, but there is some variation between them. Illumina is regarded as the leader in terms of next-generation sequencing accuracy, with the majority of bases hitting the Q30 (99.9%) benchmark. There’s no publicly available information regarding the accuracy of its long reads, though.

PacBio long-read sequencing used to be error-prone, but its latest HiFi method helped improve accuracy significantly. The average quality score for its HiFi long-read analysis is Q30, which is on par with Illumina’s short-read sequencing and a major improvement compared to previous long-read methods. Its SBB short-read sequencing has scored up to Q40 (99.99%), surpassing Illumina’s SBS method in terms of accuracy.

Source: PacBio

3. Throughput

While PacBio is known for its high accuracy scores, Illumina stands out for its efficiency and scalability. Illumina’s SBS method is designed for high throughput and can analyze millions of DNA fragments at the same time. Over the years, Illumina has also simplified the sequencing workflow and reduced the costs and prep time by enabling both short and long-read sequencing on a single platform.

Although more efficient than previous long-read methods, PacBio’s technology is relatively slow because it sequences long fragments in real time, reading bases one by one, while Illumina sequences millions of them simultaneously. The prep time is also usually longer with PacBio than Illumina, sometimes requiring as long as three days, which adds to your turnaround time.

Bonus read: Check out our comparisons of today’s popular DNA testing services:

• Ancestry vs. 23andMe

• Sequencing.com vs. Nebula

• CRI Genetics vs. 23andMe

4. Potential cost

Due to its high efficiency and economies of scale, Illumina is currently a more cost-effective option than PacBio, especially for big projects requiring a large output and fast turnaround. Illumina is also responsible for making genomics more accessible to a wider audience. The company works with numerous companies to distribute budget-friendly exome and whole-genome tests.

For instance, Illumina works with Nucleus to offer the most affordable and accessible whole-genome DNA tests in the U.S. Whole-genome sequencing (WGS) is the gold standard if you’re looking for comprehensive insights on your genetic traits and health predispositions. However, most people are unable to access WGS testing as it can cost thousands of dollars.

[X](https://x.com/nucleusgenomics/status/1777787765205135849)

Nucleus, backed by Illumina technology, offers its whole-genome test kit for only $399, which enables users to access their complete genetic data, as well as insights into disease risks, carrier status, rare genetic variants, etc.

5. Applications

Illumina’s sequencing platforms are highly versatile and can support a wide range of applications. Its proprietary short-read, high-throughput technology is fitting for large-scale projects that require efficiency and large volumes of data, such as:

• Genome-wide association studies: Studies that analyze the genomes of many people to associate gene variations with diseases.

• Transcriptomics: The analysis of cells’ RNA profiles to understand gene expression patterns and how they may contribute to diseases.

• Metagenomics: The study of genetic material from environmental samples, commonly from microbial communities.

Also, the lower cost of Illumina’s short-read sequencing makes it more suitable for analysis of individual patient or consumer genomes.

PacBio is the optimal choice when your project requires a high level of detail and accuracy. Its SBB system can detect rare sequences in diverse samples and subtle changes that may be present in few cells, which makes it suitable for applications such as CRISPR (clustered regularly interspaced short palindromic repeats) editing. 

As for PacBio’s long-read sequencing, the technology excels in areas such as:

• Detecting complex structural variants associated with genetic diseases

• Creating detailed and novel genome assemblies from scratch

• Studying DNA sequences that have repeated segments

• Exploring gene isoforms (different versions of the same gene with varying functions)

PacBio vs. Illumina: Takeaways

Comparing PacBio and Illumina is no straightforward task as the two companies are both highly respected and dominant in their respective fields. PacBio is a leader when it comes to long-read sequencing, while Illumina’s short-read method has been commonly used for over two decades and has made consumer-grade genetic testing possible. 

It’s worth noting that while PacBio’s contributions to accuracy are significant, it was Illumina’s high-throughput approach that democratized genomics and opened the door for various scientific advancements in the first place.

Because of Illumina, platforms like Nucleus (and many others) can offer affordable DNA tests and help a wide range of people access potentially life-saving health insights and improve their longevity and family planning.

[X](https://x.com/ddcaridi/status/1830031320946524274)

Get all-in-one genetic health insights with Nucleus

Nucleus provides you with more agency over your health by offering detailed and accurate insights that you can use to improve your quality of life. With a simple at-home DNA test, Nucleus will analyze ~100% of your DNA, targeting both:

• Common ancestry-informed disease risks across categories like:

  • Heart diseases

  • Cancers

  • Mental health

  • Food and diet

  • Autoimmune conditions

  • Neurology

  • Focus and energy

• Rare genetic variants (that many other tests miss) that can have a large impact on your health

Nucleus tests are physician-approved and clinical grade, which adds a layer of integrity to its analyses. Another aspect that makes Nucleus stand out is its consideration of non-genetic and lifestyle factors, like your cholesterol level and smoking status, to provide tailored risk estimates.

Source: Nucleus

Through Nucleus’ easy-to-follow reports, you can learn about:

1. Your risk for 170+ diseases and conditions, including rare cancers

2. Genetic conditions that run in your family

3. Your genetic intelligence score and other traits, such as muscle strength and longevity

By partnering with SteadyMD, Nucleus also offers genetic counseling services in case you need guidance to act on your results. 

Nucleus is HIPAA-compliant and works with fully regulated U.S. labs that use cutting-edge Illumina technology. You can download your DNA data anytime by accessing your account.

Source: Nucleus

Start your self-discovery journey with a Nucleus kit

Order your Nucleus kit today to send your sample and unlock access to all available health reports. You can also access updated reports based on research advancements and your lifestyle changes. 

To get your Nucleus kit, do this:

1. Sign up for Nucleus Premium

2. Add your personal information

3. Order the kit

Once your kit arrives at your home, swab your cheeks and send the samples to the lab. You’ll get your reports online in 6–8 weeks.

Source: Nucleus

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Featured image source: National Cancer Institute