Twisted CRISPR: A Novel Activation Strategy to Treat Genetically Driven Obesity

Two Is Better Than One

Obese and normal mouse

Redundancy equips us to survive. We have more than one lung or one kidney for a reason—if one organ in a pair gets damaged, we can still manage if the other is functional. At the cellular level, we have two copies of each chromosome in every non-germline cell. Each copy was inherited originally from a single sperm and ovum, which are “haploid” cells. Consequently, there are two copies of any given gene in non-germline “diploid” cells. In many cases, should one copy of a gene be damaged, the cell can still survive with the other, functional copy of a gene. In plants, this redundancy is common, and many plants exhibit polyploidy. In an extreme example of polyploidy, the large (by bacterial standards) but otherwise unassuming species Epulopiscium contains tens of thousands of copies of its genome.

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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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Microsatellite Instability Symposium at Duke University

On January 23, doctors, scientists and researchers will gather for a symposium about Microsatellite Instability (MSI) at Duke University. During the one-day event, scientists from Duke University and The Ohio State University will share insight into their research on biomarkers, MSI status and GI cancer, Lynch Syndrome, and MSI and DNA mismatch repair deficiency (dMMR).

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How Cheese Helped Us Understand Microbial Interactions

In almost every environment on earth, such as soil, human skin and gut, there lives a whole community of microbes—sometimes up to hundreds of species. It may seem like they all flourish in peace. But just like you may have friendly or hostile interactions with your neighbors, the different bacterial species interact in various ways. They may cooperate, compete or, sometimes, even kill each other. The interaction is complicated, and scientists have struggled to understand the nature of these microbiome interactions. How do microbiomes assemble and maintain stability? How do the interactions among different species affect gene expression?

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Combatting Gun Violence with Synthetic Biology

Imagine you are a high school student living in a community devastated by gun violence and death. In the U.S., this could be one of many communities, but it happens to be Baltimore which had 301 deaths due to gun violence in 2017 (with a per capita rate well above other large cities). Then imagine you were part of an organization within that community that helped you, along with other students, gain knowledge and skills to come up with a viable solution to the problem using synthetic biology.

Baltimore Bio-Crew at the 2018 iGEM Giant Jamboree

This is exactly how the Baltimore Bio-Crew came up with their iGEM project, Coagulance Rx. The Baltimore Bio-Crew decided to tackle this community issue head-on. One team member, Mercedes Ferandes, reflected, “Living in Baltimore City, I have not only witnessed gun violence in front of me, but have had family members and friends die from it. I wanted to try to decrease the amount of deaths by gun violence using iGEM.”

After some research, they discovered that many of the gun deaths were due to blood loss and could have been prevented. The impoverished neighborhoods where this violence occurs lack the resources to provide timely emergency medical treatment. Many of these deaths can be attributed to delayed arrival of emergency response teams—wait times for an ambulance can be over an hour.

Although there were several contributing factors beyond their control, the team wanted to address this problem by focusing on blood clotting and how it could be helpful as a quick temporary treatment for open wounds. This solution could offer a reliable, cost efficient way to save lives by slowing or stopping blood loss until a victim could get medical attention. The team decided to pursue the use of snake venom after coming across some previous iGEM projects that had used it for clotting. Team member Henry Ryles pointed out that the need for snake venom powerful enough to clot blood quickly led them to choose the venom of the Russell’s Viper
(Daboia russelii).

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A Roadmap for PROTAC Development

PROTACs or Proteolysis-Targeting Chimeras are an emerging tool in protein degradation studies, potentially suited to any need involving the removal of a specific protein. These small-molecule chimeras are exciting due to: 1) their target specificity; and 2) their ability to enable target destruction versus target inhibition.

Destruction/Inhibition: Is There a Difference?
An analogy that microbiologists (and wrestlers or anyone that has ever spent time in a locker room shower) would understand, is fungicidal versus fungistatic compounds. A fungicidal compound kills fungus. A fungistatic compound just slows the fungus down a bit.

A small-molecule inhibitor attaches to its target protein, but for how long? What inhibitor testing must be done to determine how long the inhibition lasts?

On the other hand, a small-molecule agent that causes protein degradation first targets the protein of interest, then attaches ubiquitin to that target. Once a protein is marked with ubiquitin, it’s a dead man. E3 ligase must be involved, but if the ubiquitin is added by E3, the end is near. Next stop, Hades.

This ubiquitinated protein is headed to the proteasome and proteins that go there don’t come back. Ubiquitination was called the ‘molecular kiss of death’ when this discovery was awarded the Nobel prize in Chemistry in 2004.

PROTAC components: target protein ligand, E3 ligase and linker.

About PROTACs
PROTACs are degrader molecules composed of three parts: 1) a ligand that is specific for the target protein; 2) a ligand for E3 ligase; and 3) a linker molecule that connects the two ligands. The E3 ligase is one of three enzymes that can add ubiquitin to a cellular component, but only ubiquitins added by the E3 ligase cause targeting to the proteasome (Zoppi et al.).

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Understanding Mechanisms of Pesticide Resistance to Thiamethoxam in the Cotton Aphid

A. gossypii on cotton leaf. Image credit: Clemson University - USDA Cooperative Extension Slide Series, , United States [CC BY 3.0 (https://creativecommons.org/licenses/by/3.0)], via Wikimedia Commons
A. gossypii on cotton leaf. Image credit: Clemson University – USDA Cooperative Extension Slide Series, , United States [CC BY 3.0 (https://creativecommons.org/licenses/by/3.0)], via Wikimedia Commons

The extensive and repetitive use of neonicotinoids has led to the development of resistance in several insect species including, the cotton aphid, A. gossypii. A. gossypii is a widely distributed pest that affects watermelons, cucumbers, pumpkin, cotton, and citrus crops, among others, making it one of the most economically important agricultural pests known. Thiamethoxam is a neonicotinoid insecticide that irreversibly binds to the nicotinic acetylcholine receptors (nAChRs) of cells in the nervous system and interferes with the transmission of nerve impulses in insects (1).

To further understand the mechanisms of resistence to thiamethoxam and other neonicotinoids, Wu et al. recently investigated (2) expression changes in the transcripts of P450 in thiamethoxam-susceptible and thiamethoxam-resistant cotton aphid strains. Nine P450 genes were significantly overexpressed in the resistant strain (especially CYP6CY14). The involvement of overexpressed P450s was examined through RNA interference (RNAi) introduced via artificial diet and dsRNA feeding.

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“We’re From NASA”: How Citizen Science Helped Find Ultima Thule

The science world is a-twitter with excitement lately, following the recent arrival of the New Horizons spacecraft at 2014 MU69, dubbed “Ultima Thule” by popular vote. The name means “beyond the borders of the known world”, signifying Ultima Thule’s status as the most distant object ever visited by Earthly spacecraft. Ultima Thule is a dark reddish rock in the Kuiper belt, a contact binary formed by two smaller rocks coming together in what was presumably a gentle fashion.

Do you wanna build a snowman? Credit: NASA/Johns Hopkins University Applied Physics Laboratory/Southwest Research Institute

Reaching this space snowman 6.5 billion kilometers away from Earth took brains, dedication, ingenuity and the help of an unnamed Argentinian man and his daughter.

To successfully intercept Ultima Thule, the New Horizons mission team needed to answer some questions, such as “What trajectory is Ultima Thule on?” and “Is there any space debris around Ultima Thule that will destroy our spacecraft?” Being so small (~30km diameter at its widest point), observing Ultima Thule directly from this far away would be too difficult, so the team relied on data gathered during stellar occultations, i.e., when Ultima Thule passed in front of a star.

One of these occultations occurred on July 17, 2017, in the Patagonia region of Argentina. The team had already struck out twice in trying to observe Ultima Thule passing over a star: once in South Africa, and again using the airborne telescope SOFIA over the Pacific Ocean, so tension was already running high.

On this particular night, it happened to be very windy where the observation team was, which is bad news when you’re trying to hold steady focus on a tiny object that’s really far away. The team found themselves needing help to shield the telescopes they had brought with them from wind vibrations, and get the data from the star “without it jiggling around all over the place”, as planetary scientist Anne Verbiscer puts it.

Where does one find volunteers for an astronomical observation? Well, apparently even in Argentina NASA is known and loved, and help can be found just by walking into the community. “If you just started out with ‘We’re from NASA,’ people started coming out of the woodwork,” said Dr. Verbiscer. And that is how one Argentinian man and his daughter ended up spending their evening blocking the wind from a telescope using a truck, a tarp and some plywood, allowing the NASA folks to collect the data they needed to send New Horizons to Ultima Thule.

Want to learn more about the search for Ultima Thule? Check out the episode of NOVA that inspired this blog!

2019: International Year of the Periodic Table

Periodic table of the elements

From the inside covers of elementary science textbooks to the walls of chemistry labs all around the world, the periodic table is one of the most pivotal and enduring tools of modern science. To honor the 150th anniversary of its discovery, the United Nations General Assembly and UNESCO have declared 2019 to be the International Year of the Periodic Table of Chemical Elements.

As with all scientific progress, Dmitri Mendeleev’s periodic table was the result of decades—centuries, even—of research performed by scientists all over the world. Aristotle first theorized the existence of basic building blocks of matter over 2,500 years ago, which later were believed to be earth, air, fire and water. Alchemist Hennig Brand is credited with discovering phosphorus in the late 17th century, sparking chemists to begin pursuing these basic atomic elements.

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Dane County High School Students Attend Stem Cell Conference at Wisconsin Institutes for Discovery

Dane County highschool students in attendance at the stem cell conference.


Embryonic stem cells have the extraordinary ability to divide without limit yet maintain the potential to make all types of cells found in the human body.  This holds tremendous implications for the worlds of drug discovery and testing, cell production, and tissue transplantation medicine.

Benjamin


Overall, I’m really glad I decided to go to the talk.  I got to learn a lot about stem cells, how they are used in different parts of the body, and some of the difficulties with using stem cells.  It was definitely way more enjoyable than anything else I was planning on doing during that time.  If there are more opportunities like this that come up, I would definitely try to go to them. 

Thomas

Celebrating the 20th anniversary of Dr. James Thomson’s breakthrough work with induced pluripotent stem cells, the Wisconsin Institute for Discovery (WID) hosted a panel of University of Wisconsin stem cell scientists to discuss the future of their research on November 13th.  Entitled “Stem Cell Science: The Next 20 Years” and designed for the general public, the audience heard from Drs. Lynn Allen Hoffman, David Gamm and David Vereide, who talked about applying stem cell research to develop clinical applications for skin grafting, vision restoration and regenerative biology, respectively.

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