PacBio Sequel IIe: Next-Gen DNA Sequencing Platform
The PacBio Sequel IIe system is revolutionizing DNA sequencing with its cutting-edge technology. This platform delivers highly accurate long reads, pushing the boundaries of genomic research1.
With the ability to generate HiFi reads up to 30 kb in length and accuracy surpassing 99.9%, the Sequel IIe sets a new standard in DNA sequencing1. Its SMRT Cell 8M technology offers an impressive 8X increase in data output compared to previous versions, making it a powerhouse for researchers1.
The Sequel IIe’s versatility shines through its various applications. From whole genome sequencing to metagenome analysis, RNA sequencing to epigenetics, this platform covers a wide range of research needs2. Its high-throughput capabilities and cost-effectiveness make it an attractive choice for labs seeking to push the boundaries of genomic exploration.
One of the standout features of the Sequel IIe is its ability to produce up to 4,000,000 HiFi reads with over 99% accuracy. This level of precision opens new doors in understanding complex genomes and detecting subtle variants. The system’s flexible runtime options, coupled with automated consumable handling, streamline the sequencing process for researchers.
The PacBio Sequel IIe is not just about raw power; it’s designed with user experience in mind. Its intuitive run setup and automated features make it accessible to researchers of varying expertise levels. This blend of advanced technology and user-friendliness positions the Sequel IIe as a game-changer in the field of long-read sequencing.
Key Takeaways
- Generates HiFi reads up to 30 kb with over 99.9% accuracy
- SMRT Cell 8M technology offers 8X more data output
- Supports various applications including whole genome and RNA sequencing
- Produces up to 4,000,000 HiFi reads per run
- User-friendly design with automated features
- Suitable for high-throughput, cost-effective sequencing
- Pushes boundaries in genomic research and variant detection
Overview of PacBio Sequel IIe Technology
The PacBio Sequel IIe revolutionizes DNA sequencing with its advanced capabilities. This platform introduces groundbreaking features that enhance genomic research and analysis.
Key Innovations in Sequencing
The Sequel IIe system brings 5-base genome sequencing to the forefront, enabling researchers to read A, T, G, C, and 5mC in native DNA. This innovation provides immediate access to the epigenome without special workflows. The system’s HiFi reads offer deeper biological insights and faster results, reducing file storage needs by up to 90% and cutting secondary analysis processing time by 70%3.
Advantages Over Previous Models
Compared to its predecessors, the Sequel IIe shines with improved performance. It boasts up to 3X higher throughput per SMRT Cell 8M, better accuracy, and shorter run times at a lower cost per base4. The system’s versatility is evident in its application range, from whole genome sequencing to metagenome analysis4.
HiFi reads from the Sequel IIe have proven crucial in major genomic projects, such as the Telomere-to-Telomere Consortium and Human Pangenome Reference Consortium3. The precisionFDA Truth Challenge V2 highlighted the superior precision and recall of variant calling methods using these reads, making the Sequel IIe a go-to choice for projects demanding high accuracy, long read lengths, and cost-effectiveness3.
Applications in Genomic Research
The PacBio Sequel IIe system opens new frontiers in genomic research. Its advanced capabilities revolutionize whole genome sequencing, metagenome sequencing, and RNA sequencing. This powerful tool empowers scientists to explore complex genomes and detect variants with unprecedented accuracy.
Understanding Complex Genomes
The Sequel IIe excels in decoding intricate genetic structures. It handles whole genome sequencing of humans, plants, animals, and microbes with ease. The system’s long-read technology, producing reads exceeding 10 kb with an N50 over 20 kb, allows researchers to navigate through repetitive regions that challenge other methods5.
In metagenome sequencing, the Sequel IIe shines by revealing microbial community diversity. Its ability to generate up to 1 billion bases per SMRT cell enables comprehensive analysis of complex microbial ecosystems5.
Improved Variant Detection
The Sequel IIe’s exceptional accuracy enhances variant detection across diverse cancer types. Its integration of long-read and short-read sequencing capabilities allows for in-depth analysis of cancer genomes at the single-cell level6. This breakthrough facilitates the discovery of clinically relevant mutations, particularly in cancers prevalent in Asia.
RNA sequencing on the Sequel IIe provides deep insights into cDNA sequences. The platform’s prowess in detecting gene isoforms and discovering novel genes pushes the boundaries of transcriptomics research5. This capability is crucial for understanding gene expression patterns in complex diseases.
| Application | Key Advantage | Impact on Research |
|---|---|---|
| Whole Genome Sequencing | Long reads (N50 > 20 kb) | Resolves complex genomic regions |
| Metagenome Sequencing | High throughput (up to 1 billion bases/cell) | Comprehensive microbial diversity analysis |
| RNA Sequencing | Accurate isoform detection | Enhanced transcriptome characterization |
Comparison with Other Sequencing Technologies
The DNA sequencing landscape offers diverse technologies, each with unique strengths. Let’s explore how PacBio’s long-read sequencing compares to other popular platforms.
PacBio vs. Illumina
PacBio’s long-read sequencing excels in handling complex genomes, while Illumina’s short-read sequencing is ideal for high-throughput needs. PacBio HiFi sequencing typically achieves over 99.9% accuracy, making it perfect for applications demanding high-quality base calling7. With read lengths of 15,000 to 20,000 bases or more, PacBio HiFi can span large genomic regions and repeats effectively7.
PacBio vs. Oxford Nanopore
Both PacBio and Oxford Nanopore Technologies (ONT) offer long-read sequencing, but with distinct differences. ONT systems provide reads ranging from 20 to over 4 Mb in length, surpassing PacBio in maximum read length7. PacBio HiFi systems typically yield 90 Gb per cell, while ONT systems deliver 50-100 Gb7.
| Feature | PacBio HiFi | Oxford Nanopore |
|---|---|---|
| Sequencing accuracy | >99.9% | >10% error rate (single pass) |
| Read length | 15,000-20,000+ bases | 20 to 4M+ bases |
| Methylation calling | On-instrument | Off-instrument |
| Storage costs (monthly) | ~$1.30 USD | ~$30.00 USD |
PacBio Sequel II assemblies show the highest consensus accuracy among tested organisms, even after adjusting for sequencing throughput differences8. Both technologies can identify SNVs and SVs, but PacBio HiFi also excels in calling Indels7. The choice between these platforms depends on project goals, budget, and specific research requirements.
Sample Preparation and Workflow
The PacBio Sequel IIe system revolutionizes DNA sequencing with its streamlined library preparation and efficient sequencing workflow. This advanced platform integrates automated consumable handling and intuitive software, simplifying the entire process from sample to results.
Steps in Sample Preparation
Proper sample preparation is crucial for optimal sequencing results. DNA samples should have a purity of OD 260/280 between 1.8 and 2.0, with OD 260/230 greater than 2.09. For best results, submit DNA in EB buffer or TLE buffer with specific pH requirements9.
The library preparation process involves several key steps:
- DNA extraction and purification
- Quality assessment
- Fragmentation (if needed)
- End repair and adapter ligation
- Size selection
- Quality control
Recommended Protocols
PacBio offers various protocols tailored to different sample types and research goals. For microbial DNA samples, silica spin column protocols are recommended9. These methods typically yield bacterial DNA fragments of about 15 to 20 kb in size, ideal for SMRT Cell loading9.
The sequencing workflow on the Sequel IIe is fully automated, eliminating manual interventions during runs. As each SMRT Cell completes, data becomes available for immediate analysis, enhancing overall efficiency10. This streamlined process can generate up to 4 million 99.9% accurate reads in under 30 hours11.
| Feature | Specification |
|---|---|
| Read Length | 15-18 kb (HiFi reads) |
| Read Quality | Q30 or better |
| Data Yield | 60-90 Gb |
| Run Time | Under 30 hours |
By following these protocols and leveraging the Sequel IIe’s advanced features, researchers can achieve high-quality sequencing results efficiently and reliably.
Data Analysis Capabilities
The PacBio Sequel IIe system brings powerful data analysis capabilities to genomic research. This advanced platform integrates cutting-edge bioinformatics tools with efficient data processing methods, revolutionizing how scientists interpret sequencing results.
Software Tools for Analysis
At the heart of the Sequel IIe’s analysis capabilities is SMRT Link, a comprehensive software suite designed for PacBio sequencing data. SMRT Link offers a user-friendly interface for managing sequencing runs, data analysis, and result visualization. The system’s on-board analysis capabilities are a game-changer for labs without high-performance computing access12.
SMRT Link v11.1 streamlines data processing, delivering HiFi reads with methylation calls directly. This feature simplifies epigenetic studies and reduces file storage needs by up to 90%3. The software also cuts secondary analysis processing time by 70%, allowing researchers to move from raw data to actionable insights faster than ever3.
Interpretation of Results
The Sequel IIe system excels in result interpretation across various applications. Its high-accuracy HiFi reads enable precise variant calling, achieving top scores in the precisionFDA Truth Challenge V2 for human genome analysis3. This accuracy extends to whole genome sequencing, targeted sequencing, and metagenomics studies13.
For RNA sequencing, the system can sequence full-length mRNA transcripts, providing a complete picture of gene expression13. In epigenetics, the 5-base sequencing capability allows researchers to explore epigenomes seamlessly, detecting methylation patterns across the genome13.
| Application | Key Benefit | Analysis Feature |
|---|---|---|
| Whole Genome Sequencing | Contiguous assemblies | SMRT Link Genome Assembly |
| Epigenetics | Methylation detection | 5-base HiFi sequencing |
| RNA Sequencing | Full-length transcripts | IsoSeq analysis |
| Variant Calling | High precision and recall | HiFi read analysis |
The Sequel IIe’s data analysis capabilities make it a versatile tool for genomic research. Its combination of advanced software and efficient data processing empowers scientists to unlock new insights from complex genomic data.
Real-World Case Studies
The PacBio Sequel IIe system has revolutionized genomic research and sequencing applications. This powerful tool has enabled scientists to make groundbreaking discoveries and push the boundaries of genetic understanding.
Research Breakthroughs Using Sequel IIe
The Sequel IIe system has significantly improved the efficiency of genomic research. It delivers a 70% reduction in overall secondary analysis time, depending on the application14. This speed boost allows researchers to process more data in less time, accelerating scientific discoveries.
One of the most impressive features of the Sequel IIe is its ability to provide HiFi reads. These reads combine the accuracy of Sanger sequencing (>99.9%) with long reads up to 25 kb14. This high accuracy enables researchers to detect both single nucleotide and structural variants simultaneously, advancing human genetics research14.
Industry Adoption Examples
The Sequel IIe has been widely adopted across various industries. In the healthcare sector, PacBio partnered with Prenetics to expand the utility of next-generation sequencing in both consumer and clinical settings15. This collaboration aims to improve the accuracy of detecting mutations in carrier screening and cancer-risk screening15.
The whole genome sequencing capabilities of the Sequel IIe have also found applications in consumer-oriented products. Prenetics is using PacBio technology for various applications, including aging, skincare, and population health studies15.
| Application | Benefit |
|---|---|
| Human Genetics Research | Accurate detection of genetic variants |
| Cancer Screening | Improved mutation detection |
| Consumer Genetics | Enhanced insights into aging and skincare |
| Population Health | Comprehensive genetic data for large-scale studies |
The impact of the Sequel IIe extends beyond healthcare. In biomedical research, the system’s ability to provide a comprehensive view of genomes, transcriptomes, and epigenomes has led to its use in thousands of peer-reviewed publications15. Scientists globally are leveraging this technology to drive discoveries in plant and animal sciences, and microbiology15.
Benefits of Using PacBio Sequel IIe
The PacBio Sequel IIe system brings significant advantages to genomic research. This advanced platform combines HiFi accuracy with long-read sequencing to enhance research efficiency.
High Accuracy and Read Length
The Sequel IIe delivers exceptional accuracy with HiFi reads surpassing 99% precision and read lengths of 15-20 kb16. This high-quality data output enables researchers to explore complex genomic regions with confidence. The system can generate up to 4,000,000 HiFi reads per run, providing a wealth of data for analysis16.
Cost-Effectiveness for Research
Despite its advanced capabilities, the Sequel IIe offers cost-effective sequencing. It produces up to 30 Gb of data per run, representing a tenfold increase in yield compared to its predecessor1716. This improved throughput translates to lower costs per base, making large-scale genomic projects more feasible for research institutions.
User-Friendly Experience
The Sequel IIe is designed for ease of use. With a run time of up to 30 hours, it offers flexibility for various research needs16. The system’s automated processes and intuitive setup reduce hands-on time, allowing researchers to focus on data analysis rather than instrument operation.
| Feature | Specification |
|---|---|
| Read Length | 1x20000bp |
| Data Quality | >99% accuracy |
| Output per Run | Up to 30 Gb |
| Run Time | Up to 30 hours |
The PacBio Sequel IIe’s combination of accuracy, efficiency, and user-friendliness has led to its rapid adoption. Over 75 systems have been installed worldwide, with the total data generated already surpassing that of all previously installed Sequel systems over four years17. This widespread use underscores the system’s value in advancing genomic research across various fields.
Limitations and Considerations
The PacBio Sequel IIe, while advanced, faces sequencing challenges. Researchers must weigh the initial investment against project needs. The system’s complexity can lead to a steep learning curve, impacting implementation timelines.
Challenges in Implementation
Long-read sequencing technologies like PacBio and Oxford Nanopore (ONT) have unique strengths, but also limitations. PacBio CLR and ONT produce longer reads than PacBio RS II and HiFi, offering better genome contiguity18. However, both have error rates exceeding 10% for single-pass sequencing18. This highlights the technology limitations in accuracy for certain applications.
Areas for Improvement
Future improvements should focus on reducing costs and expanding applications. Long-read sequencing libraries are pricier than short-read alternatives18. Enhancing throughput and lowering expenses could make the technology more accessible. The Illumina genome sequencing approach offers insights into cost-effective strategies that could be adapted.
Addressing chimeric reads, present in both PacBio and ONT workflows, is crucial18. Improving read accuracy and reducing error rates below the current 1% for PacBio HiFi would broaden the technology’s applicability19. These enhancements could particularly benefit projects like the All of Us initiative, which aims to sequence over one million diverse Americans19.
| Technology | Strengths | Limitations |
|---|---|---|
| PacBio Sequel IIe | High accuracy (HiFi mode), long reads | High cost, complex implementation |
| Oxford Nanopore | Ultra-long reads (up to 4 Mbp) | Higher error rate |
| Illumina | Cost-effective, high throughput | Shorter read lengths |
Future of DNA Sequencing Technology
DNA sequencing is evolving rapidly, with long-read technologies like PacBio HiFi sequencing gaining popularity in genomics research20. These sequencing innovations are reshaping the landscape of genomic technology advancements, promising exciting developments in the coming years.
Upcoming Features and Enhancements
PacBio’s HiFi sequencing produces reads 15,000 to 20,000 bases long with 99.9% accuracy, earning it the title of 2022 Method of the Year by Nature Methods20. This technology allows for epigenetic studies by measuring base incorporation speed, offering new insights into genetic regulation20.
Future enhancements in sequencing technology are likely to focus on increasing accuracy, read length, and throughput while reducing costs. The SMRT Link software used to analyze HiFi sequencing data is continuously improving, aiming to streamline data interpretation and analysis20.
Potential Market Impact
The sequencing market is dynamic, with various players competing for dominance. While Illumina currently generates 90% of the world’s sequence data, other technologies are making strides21. Nanopore PromethION leads in output per hour, followed by BGI and Illumina21.
Market trends indicate a shift towards more affordable and efficient sequencing methods. The cost of sequencing a human genome has dropped dramatically from the initial $2.7 billion to just $10 million, showcasing the rapid pace of innovation in this field21. As these technologies continue to advance, we can expect further reductions in cost and increases in accessibility, potentially revolutionizing personalized medicine and biotechnology industries.
| Sequencing Technology | Read Length | Accuracy | Key Advantage |
|---|---|---|---|
| PacBio HiFi | 15,000-20,000 bases | 99.9% | Long reads with high accuracy |
| Illumina | 50-300 bases | High | Market dominance |
| Nanopore PromethION | Variable | Improving | Highest output per hour |
Customer Support and Resources
PacBio provides extensive support for Sequel IIe users, ensuring researchers can maximize their system’s potential. The company offers a range of resources to assist with various aspects of sequencing projects.
Getting Help with Your Sequel IIe
When you need technical support for your Sequel IIe, PacBio’s expert team is ready to assist. They offer guidance on system operation, troubleshooting, and best practices. The SMRT Link User Guide, updated regularly, covers the Sequel IIe system comprehensively, helping users navigate common challenges10.
Online Resources and Training
PacBio’s online platform is a treasure trove of sequencing resources. It features user guides, application notes, and webinars tailored to Sequel IIe users. The platform provides detailed information on HiFi application options specific to the Sequel IIe system, enabling researchers to optimize their workflows10.
Training programs are a key component of PacBio’s support strategy. These programs cover various topics, from sample preparation to data analysis. For instance, PacBio offers guides on using the Nanobind CBB Kit for DNA extraction from different sample types, specifically designed for Sequel IIe users10. This hands-on approach ensures users can fully leverage the system’s capabilities.
By providing comprehensive technical support, rich sequencing resources, and targeted training programs, PacBio empowers Sequel IIe users to push the boundaries of genomic research.
Conclusion: Embracing Next-Gen Sequencing
The PacBio Sequel IIe system has revolutionized genomic research future. Since PacBio’s founding in 2005, the company has made significant strides in sequencing technology impact. The Sequel IIe, launched to boost efficiency and accuracy, can now achieve up to 99.999% accuracy for genome assembly and variant detection22.
The Role of Sequel IIe in Future Research
Sequel IIe’s long-read capabilities, with average lengths of 8-15kb and peaks of 40-70kb, open new doors for complex genome studies. Its ability to detect minor variants at 0.1% frequency and directly identify base modifications enhances our understanding of genetic variations22. This technology’s minimal GC bias ensures uniform coverage, addressing limitations faced by other platforms like Illumina2223.
Encouraging Innovation in Genomics
The Sequel IIe’s impact extends beyond basic research. Its applications in vision research have doubled NGS-based studies in the eye field since 201524. The system’s accuracy and long reads have facilitated the identification of 240 genes linked to retinal degenerative diseases, showcasing its potential in uncovering rare variants in complex diseases24.
As we look to the future, the PacBio Sequel IIe stands at the forefront of scientific innovation. Its unique features and growing adoption in various fields promise to drive genomic research forward, paving the way for breakthroughs in personalized medicine and our understanding of complex biological systems.
Q&A
What is the PacBio Sequel IIe system?
The PacBio Sequel IIe system is an advanced DNA sequencing platform that delivers highly accurate long reads. It offers up to 4,000,000 HiFi reads with >99% accuracy and uses SMRT Cell 8M technology. The system is designed for high-throughput, cost-effective sequencing across various applications including whole genome sequencing, metagenome sequencing, RNA sequencing, epigenetics, and targeted sequencing.
What are the key innovations of the Sequel IIe system?
The Sequel IIe system introduces 5-base genome sequencing (A, T, G, C, 5mC) in native DNA, providing immediate access to the epigenome without special workflows. It offers deeper biological insights, less data processing, and faster results due to the clarity of HiFi reads. Compared to previous models, it provides higher throughput, increased accuracy, and improved cost-effectiveness.
What applications does the Sequel IIe system support?
The Sequel IIe system supports various applications in genomic research, including whole genome sequencing for humans, plants, animals, and microbes, metagenome sequencing to reveal microbial community diversity, RNA sequencing for in-depth analysis of cDNA sequences, and targeted sequencing to study specific genome regions of interest.
How does PacBio’s long-read sequencing compare to other technologies?
PacBio’s long-read sequencing offers high accuracy and works well for complex genomes, while short-read sequencing (like Illumina) is cost-effective for high-throughput needs. PacBio’s HiFi reads provide exceptional clarity and accuracy compared to Oxford Nanopore’s long reads. The choice between technologies depends on factors such as project goals, budget, accuracy requirements, and timeline.
What are the sample preparation and workflow features of the Sequel IIe system?
The Sequel IIe system features simplified, unified, and accelerated library preparation workflows and consumables. It offers automated consumable handling with integrated software and intuitive run setup. The sequencing process is fully automated with no manual touch points during a run, and data is available for analysis as each SMRT Cell completes.
What software does the Sequel IIe system use for data analysis?
The Sequel IIe system uses SMRT Link v11.1 for data analysis. It offers direct output of HiFi reads with methylation calls, simplifying epigenetic studies. The system’s software capabilities allow for efficient data processing and interpretation, providing clear results across various applications.
What are the benefits of using the PacBio Sequel IIe system?
The Sequel IIe system offers high accuracy with HiFi reads >99% accurate and long read lengths. It provides cost-effective high-throughput sequencing for various applications. The user-friendly experience includes automated processes, intuitive setup, and flexible runtime options. The system’s ability to perform 5-base genome sequencing without additional workflows enhances research efficiency and cost-effectiveness.
Are there any limitations or considerations when using the Sequel IIe system?
While the Sequel IIe system offers numerous advantages, researchers should consider factors such as initial investment, specific project requirements, and potential learning curves when implementing the technology. Areas for potential improvement may include further increasing throughput, reducing costs, and expanding application ranges.
What support and resources are available for Sequel IIe users?
PacBio offers comprehensive customer support for Sequel IIe users. This includes technical assistance, online resources, and training programs. Customers can connect with PacBio scientists for expert advice and access a wealth of online materials including user guides, application notes, and webinars to maximize their use of the Sequel IIe system.
What is the future outlook for the PacBio Sequel IIe system?
PacBio continues to innovate with regular software and hardware updates. Future enhancements may focus on increasing accuracy, read length, and throughput while reducing costs. The advancement of long-read sequencing technology is expected to have significant impacts on genomic research, personalized medicine, and biotechnology industries.
