NGS vs Sanger
NGS vs Sanger sequencing
Understanding the key differences and when next-generation sequencing can be a more effective option than Sanger sequencing
Differences between NGS and Sanger sequencing
In principle, the concepts behind Sanger vs next-generation sequencing (NGS) technologies are similar. In both NGS and Sanger sequencing (also known as dideoxy or capillary electrophoresis sequencing), DNA polymerase adds fluorescent nucleotides one by one onto a growing DNA template strand. Each incorporated nucleotide is identified by its fluorescent tag.
The critical difference between Sanger sequencing and NGS is sequencing volume. While the Sanger method only sequences a single DNA fragment at a time, NGS is massively parallel, that is, sequencing millions of fragments simultaneously per run. This process translates into sequencing hundreds to thousands of genes at one time. NGS also offers greater discovery power to detect novel or rare variants with deep sequencing.
Advantages of NGS vs Sanger sequencing
Advantages of NGS include:
- Screen more samples cost-effectively
- Detect multiple variants across regions of the transcriptome
- Increase sequencing depth, mutation resolution/sensitivity and discovery power
- Higher sensitivity to detect low-frequency variants1, 2
- Faster turnaround time for high sample volumes3
- Comprehensive genomic coverage
- Lower limit of detection4, 5
- Higher capacity with sample multiplexing
- Ability to sequence hundreds to thousands of genes or gene regions simultaneously
Sanger sequencing and NGS comparison
| Sanger sequencing Sanger sequencing interrogates a gene of interest. |
Targeted NGS Targeted NGS simultaneously sequences several hundreds to thousands of genes. |
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| Benefits1-7 |
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| Challenges1-7 |
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* Discovery power is the ability to identify novel variants.
† Mutation resolution is the size of the mutation identified. NGS can identify large chromosomal rearrangements down to single nucleotide variants.
Using NGS instead of Sanger sequencing
Watch this animation to see how the easy and accessible Illumina NGS technology can complement your Sanger sequencing work.
"With Sanger sequencing, we saw a limited DNA snapshot of each sample, typically obtaining 50–100 reads per sample. NGS and its massively parallel sequencing enable us to look at tens to hundreds of thousands of reads per sample."
When to use NGS vs Sanger sequencing
Sanger sequencing can be a good choice when interrogating a small region of DNA on a limited number of samples or genomic targets (~20 or fewer). Otherwise, targeted NGS is more likely to suit your needs. NGS allows you to screen more samples cost-effectively and detect multiple variants across targeted areas of the genome—an approach that would be costly and time-consuming using Sanger sequencing.
Getting started with NGS
For an in-depth look at how NGS compares to other methods, download our Getting Started with NGS eBook. This 20+ page eBook outlines key differences, recommended methods/applications, sample workflows, and more.
Download eBook
NGS applications and methods
No matter your research focus, NGS can play an important role in pursuing the answer to a variety of biological questions using a wide array of published methods for diverse sample types. With hypothesis-free experimental design, NGS is unearthing new knowledge on cancer, microbiology, genetic disease, reproductive health, agriculture, and more.
Further reading
Benchtop sequencers
Most researchers start with an in-lab benchtop sequencing system. This resource is a great place to learn about benchtop sequencing capabilities, instruments, and popular applications and methods for each.
NGS vs qPCR
While qPCR is effective for low target numbers, the workflow can be cumbersome for multiple targets. NGS is preferable for studies with many targets or samples. Learn about the differences and benefits of each technology.
NGS for beginners
Curious how NGS can benefit your research goals? This page offers simple, clear explanations of next-generation sequencing and its numerous benefits over conventional methods.
Using Sanger but curious about NGS?
If you have questions about NGS for your specific research focus, we’d love to help. Our specialists can answer any questions and recommend the best solution for your setup.
References
- Jamuar SS, Lam AT, Kircher M, et al. Somatic mutations in cerebral cortical malformations. N Engl J Med. 2014;371(8):733-743.
- Rivas MA, Beaudoin M, Gardet A, et al. Deep resequencing of GWAS loci identifies independent low-frequency variants associated with inflammatory bowel disease. Nat Genet. 2011;43(11):1066-1073.
- König K, Peifer M, Fassunke J, et al. Implementation of amplicon parallel sequencing leads to improvement of diagnosis and therapy of lung cancer patients. J Thorac Oncol. 2015;10(7):1049-1057.
- Shendure J and Ji H. Next-generation DNA sequencing. Nat Biotechnol. 2008;26(10):1135-1145.
- Schuster SC. Next-generation sequencing transforms today’s biology. Nat Methods. 2008;5(1):16-18.
- Myllykangas S and Hanlee JP. Targeted deep resequencing of the human cancer genome using next-generation technologies. Biotechnol Genet Eng Rev. 2010; 27: 135–158.
- Illumina. High-impact discovery through gene expression and regulation research. https://www.illumina. com/content/dam/illumina-marketing/documents/gated/gene-expression-profiling-e-book-web.pdf. Accessed March 23, 2023.