Deep in the Jungle Something Is Happening: DNA Sequencing

This blog was written by guest blogger and 2018 Promega Social Media Intern Logan Godfrey.

Only 30 years ago, the polymerase chain reaction (PCR) was used for the first time, allowing the exponential amplification of a specific DNA segment. A small amount of DNA could now be replicated until there was enough of it to study accurately, even allowing sequencing of the amplified DNA. This was a massive breakthrough that produced immediate effects in the fields of forensics and life science research. Since these technologies were first introduced however, the molecular biology research laboratory has been the sole domain of PCR and DNA sequencing.

While an amazing revolution, application of a technology such as DNA sequencing is limited by the size and cost of DNA sequencers, which in turn restricts accessibility. However, recent breakthroughs are allowing DNA sequencing to take place in jungles, the arctic, and even space—giving science the opportunity to reach further, faster than ever before. 

Gideon Erkenswick begins extractions on fecal samples collected from wild tamarins in 2017. Location: The GreenLab, Inkaterra.

Gideon Erkenswick begins extractions on fecal samples collected from wild tamarins in 2017. Location: The GreenLab, Inkaterra. Photo credit: Field Projects International.

The newfound accessibility of DNA sequencing means a marriage between fields of science that were previously largely unacquainted. The disciplines of genomics and wildlife biology/ecology have largely progressed independently. Wildlife biology is practiced in the field through observations and macro-level assessments, and genomics, largely, has developed in a lab setting. Leading the charge in the convergence of wildlife biology and genomics is Field Projects International. 

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Where Would DNA Sequencing Be Without Leroy Hood?

There have been many changes in sequencing technology over the course of my scientific career. In one of the research labs I rotated in as a graduate student, I assisted a third-year grad student with a manual radioactive sequencing gel because, I was told, “every student should run at least one in their career”. My first job after graduate school was as a research assistant in a lab that sequenced bacterial genomes. While I was the one creating shotgun libraries for the DNA sequencing pipeline, the sequencing reaction was performed using dideoxynucleotides labeled with fluorescent dyes and amplified in thermal cyclers. The resulting fragments were separated by manual loading on tall slab polyacrylamide gels (Applied Biosystems ABI 377s) or, once the lab got them running, capillary electrophoresis of four 96-well plates at a time (ABI 3700s).

Sequencing throughput has only increased since I left the lab. This was accomplished by increasing well density in a plate and number of capillaries for use in capillary electrophoresis, but more importantly, with the advent of the short read, massively parallel next-generation sequencing method. The next-gen or NGS technique decreased the time needed to sequence because many sequences were determined at the same time, significantly accelerating sequencing capacity. Instruments have also decreased in size as well as the price per base pair, a measurement used when I was in the lab. The long-prophesized threshold of $1,000 per genome has arrived. And now, according to a recent tweet from a Nanopore conference, you can add a sequencing module to your mobile device:

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Optimizing a DNA Methylation Analysis Workflow

methyledge seminarWhen Aristotle compared epigenetics to a net (1), he could not have predicted how right he was.  Recent research has revealed that mechanisms underlying epigenetic effects are numerous and interdependent as are the knots in a net. Each epigenetic mechanism has its players: enzymes, functional groups, substrates etc.  The most important aspect of an epigenetic trait is its reversibility. Methylation of DNA was the first epigenetic modification to be discovered, and 5-cytosine methylation was the first to be linked with gene expression status. Currently, the most popular method for measuring  CpG island methylation status is a bisulfite treatment of DNA followed by PCR or sequencing.

In this week’s webinar, Promega R&D scientist, Karen Reece focused on a workflow from DNA purification to analysis. She described the best methods for DNA isolation, quantification, bisulfite conversion, PCR and sequencing. Continue reading “Optimizing a DNA Methylation Analysis Workflow”

The Ongoing Legacy of the Human Genome Sequence

When the first draft sequence of the human genome was announced, I was a research assistant for a lab that was part of the Genome Center of Wisconsin where I created shotgun libraries of bacterial genomes for sequencing. Of course, the local news organizations were all abuzz with the news and sought opinions on what this meant for the future, including that of the lab’s PI and oddly enough, my own. While I do not recall the exact words I offered on camera, I believe they were something along the lines of this is only the first step toward the future of human genetics. Ten years later, we have not fulfilled the potential of the grandiose words used to report the first draft sequence but have gained enough knowledge of what our genome holds to only intrigue scientists even more.

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