How scientists decipher COVID-19 —Part 2: Sanger and Next Generation Sequencing
Next Generation sequencing (NGS) is a vast improvement upon traditional sequencing methods such that the sequencing of the human genome can be shortened from ten years to one day. Despite the popularization, NGS has yet to be translated into routine clinical practice, and is currently only used for research purposes.
How NGS works
NGS is able to sequence fragments of DNA in parallel, unlike how Sanger sequences fragments one at a time.
Then bioinformatics analyses would piece the fragments together by referring to a database.
The fragments are sequenced multiple times for accuracy and enhanced coverage. NGS can sequence whole or parts of the genome.
Why NGS is more preferable than Sanger?
1. Capture a broader spectrum of mutations.
Sanger: can only capture substitution, small insertions and deletions. Additional mutations would require further testings.
NGS: can capture all modifications (exceptions: extreme guanine/cytosine content or repeated architecture, such as Fragile X syndrome and Huntington Disease).
2. Genomes are interrogated objectively.
Sanger: depends on preknowledge of the gene.
NGS: Discover new mutations and disease causing genes in unexplained syndromes, such as unexplained developmental delay in children.
3. Allow detection of mosaic mutations.
Sanger: sequencing sensitivity is too low.
NGS: is sensitive enough to detect the mutation, and it can further increase sensitivity by increasing sequencing depth (ex. to understand foetal DNA from maternal blood, and tracking tumor cell level from patient circulation).
4. Determine the identity of pathogens through genomic data (microbiology)
Sanger: traditional methods are too time consuming. Clinical practice relies on culture dependent assays.
NGS: can determine a pathogen’s genetic composition, such as:
1) Determine identity.
2) Reveal potential drug sensitivity information.
3) Discover pathogens’ evolutionary relationship with each other, meaning you can trace sources of infection outbreak and potentially trace it back to a single staff member.
Conventional way: Morphing pathogens, getting staining properties and metabolic criteria.
5. Advance cancer genome research (oncology)
Sanger: too time intensive which results in few samples and genes of somatic mutations (mutations that cause cancer) to study from.
NGS: enables the study of the cancer genome in its entirety, and has the following benefits:
a) More precise diagnosis.
b) More precise classification.
c) More accurate prognosis.
d) Identification of drug-able causal mutations.
e) Personalized cancer management becomes possible.
f) Identify mutations in tumors.
g) Target mutations by mutation specific drugs.
There are two main constraints that prevent people from adopting the technology, they are economic constraints and technical constraints.
- Getting the required computer capacity and storage can get very expensive due to the need to store and process large amounts of data.
- Hiring needed expertise to operate can be costly.
- It is difficult to find people that are skilled that performing important data extraction.
However, IDseq, that is backed by Chan Zuckerberg Initiative, can overcome these limitations for use in places that lack resources to conduct comparable research and studies.
Special thanks to Phoenix Yin (Microbiology@UBC | Pre-med student) for answering all my questions and proofreading this article.