3. November 2020 Britta Baur

One ‘Pore To Rule Them All

© Bruce777

Origin of the Sequence

The year is 2001 and Canadian rock band, Nickelback, has released their widely acclaimed single, “How You Remind Me”. Earlier that year, a consortium of international researchers published the results of the Human Genome Project (HGP), giving us a glimpse into genetic sequences for the first time. The instrument used to accomplish this was the Sanger DNA Sequencer, a machine with roughly the same dimensions as a refrigerator and capable of producing up to 0.01 GB (roughly the size of two songs) of data per day! This technology would allow for 90% of the human genome to be sequenced over the course of a few years. In total, the HGP cost ~USD3 billion to sequence the genome of a single individual. At the time, this collaborative effort was groundbreaking, and not because the sequencer was so heavy.

Third Generation Sequencing Technology

Gibbs Kuguru Standing in front of a Sanger Sequencer (© G. Kuguru)

Fast-forward 20 years later, in the throes of a global pandemic, Oxford Nanopore Technologies (ONT) is sending DNA Sequencers by the box-load to nations around the globe to rapidly identify and monitor the SARS-Cov-2 plague. The information garnered from this endeavor will support virologists in curbing the spread of the COVID-19 disease, and can serve those in even the most remote areas. The device used to accomplish this can fit in the palm of your hand and it costs a bit more than the newest iPhone. These Nanopore sequencers are capable of producing 30 GB of data in real time, allowing for the human genome to be sequenced in a few hours! Not only is this fast, but this tool also better suited sequencing big genomes unlike previous iterations of DNA sequencers. Due to their complexity, big genomes have been extremely difficult to characterize, and the technique behind Nanopore sequencing essentially solves that problem by making the genome assembly process much easier…but actually, what does this have to do with sharks?

How big is a shark genome?

Compared to humans, the genomes of sharks can be up to twice the size! Paired with the high sequencing associated costs, decoding the genome of sharks was simply not a priority. However, the genome of the great white shark was characterized in 2019 and the insights garnered from that study were very compelling, even for biomedical research! In particular, the great white shark genes associated with adaptive healing could be used to stem the tide of cancerous tissues from developing, which may prove useful in controlling this highly volatile disease. But still, even with all of this information, we have trench-deep uncertainties as to how DNA actually works. Well, “How,” you might ask, “do we get to the bottom of this?” One method is to compare shark genomes to each other to find out which genes are crucial to the function of these healing properties…this is a field called comparative genomics. The currency of comparative genomics is genomes, and the more we have, the richer our gene banks (more like gene libraries) will be. Indeed, this convention goes for all living things. Attaining this information has never been closer to our reach than now, especially with the advent of Nanopore technologies. But wait, it definitely gets better.

 

Built Nanopore Tough

DNA is comprised of four molecules called nucleotides, which code for the physical make-up of every living being and ONT has developed a technique to rapidly read this code. By using a current, a single strand of DNA (ssDNA) travels through a tiny hole called a “pore”, and the Nanopore sequencer measures the change in resistance of the pore when its conductive channel is partially blocked by the ssDNA molecule. Since every nucleotide has it’s own electrical resistance to the pore, we can determine the sequence based on the changes in electricity per nucleotide. This technique is so robust, that conducting DNA sequencing within a laboratory environment is no longer required. To date, Nanopore sequencers have performed while strapped to researchers seeking microbial communities in the Antarctic Circle, in some of the most arduous conditions on the planet. This tech has also been used on the International Space Station in a microgravity environment with optimal success in obtaining RNA molecules! Not only does this demonstrate the ability for this tool to be used outside of the lab, but it also opens up a new world of research questions that we are now able to answer.

 

Gibbs Kuguru Holding a Nanopore Sequencer (© G. Kuguru)

I will be using Nanopore sequencing and comparative genomics to reveal the cracks in the genome of blacktip reef sharks afflicted with Leucism (https://www.sharkproject.org/en/sharks-adapting-to-humanity/). After sampling these individuals, the sequencing will be orchestrated on a beach research station on Kihaaddhufaru Island hosted by Ocean Dimensions and Kihaa Maldives. This research is funded by the Africa Talent Program from Wageningen University & Research and by Sharkproject Switzerland. Though the technological advancements supporting this study are remarkable, the challenges in this scientific endeavor will be met through a generous collaborative effort in both the private and public sector. Watch this space for more updates!

 

Credit Text & Photos: Gibbs Kuguru

Sharkproject Switzerland will initially support Gibbs Kuguru’s research project and doctoral thesis for four years. He will give regular updates on his project ‘Inbreeding and the Unexpected Presence of Leucism in Caracharhinus melanopterus’.