38 Years After First Release, RNasin Protects COVID-19 Tests

A protein first purified and sold by Promega almost four decades ago has emerged as a crucial tool in many COVID-19 testing workflows. RNasin® Ribonuclease Inhibitor was first released in 1982, only four years after the company was started. At that time, the entire Promega catalog fit on a single sheet of 8.5 × 11” paper, and RNasin was one of the first products to draw widespread attention to Promega. Today, the demand for this foundational product has skyrocketed as it supports labs responding to the COVID-19 pandemic.

What is RNasin® Ribonuclease Inhibitor?

RNA is notoriously vulnerable to contamination by RNases. These enzymes degrade RNA by breaking the phosphodiester bonds forming the backbone of the molecule. To say that RNases are everywhere is barely an exaggeration – almost every known organism produces some form of RNase, and they’re commonly found in all kinds of biological samples. They’re easily introduced into experimental systems, since even human skin secretes a form of RNase. Once they’re present, it’s very hard to get rid of them. Even an autoclave can’t inactivate RNases; the enzymes will refold and retain much of their original activity.

RNasin® Ribonuclease Inhibitor is a protein that has been shown to inhibit many common contaminating RNases, but without disrupting the activity of enzymes like reverse transcriptase that may be essential to an experiment. It works by binding to the RNase enzyme, prevent it from acting on RNA molecules. This is important for ensuring that RNA samples are intact before performing a complex assay.

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RT-qPCR and qPCR Assays—Detecting Viruses and Beyond

We have all been hearing a lot about RT-PCR, rRT-PCR and RT-qPCR lately, and for good reason. Real-Time Reverse Transcriptase Polymerase Chain Reaction (rRT-PCR) is the technique used in by the Center for Disease Control (CDC) to test for COVID-19. Real-time RT-PCR, or quantitative RT-PCR (RT-qPCR)*, is a specialized PCR technique that visualizes amplification of the target sequesnce as it happens (in real time) and allows you to measure the amount of starting target material in your reaction. You can read more about the basics of this technique, and watch a webinar here. For more about RT-PCR for COVID-19 testing, read this blog.

Both qPCR and RT-qPCR are powerful tools for scientist to have at their disposal. These fundamental techniques are used to study biological process in a wide range of areas. Over the decades, Promega has supported researchers with RT-qPCR and qPCR reagents and systems to study everything from from diseases like COVID-19 and cancer to viruses in elephants and the circadian rhythm of krill.  

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What We Know About the 2019 Novel Coronavirus, SARS-CoV-2

David Goodsell image of SARS-2-CoV
Image by David Goodsell

The 2019 Novel Coronavirus (SARS-CoV-2) is a new virus that emerged in China in late 2019 and quickly jumped into scientific and mainstream news. When facing a potential pandemic, it can be difficult to share information without inducing panic. There’s no doubt that SARS-CoV-2 presents a significant threat to public health, but as with all viruses in their emerging stages, we often find ourselves with more questions than answers. However, through the work of the World Health Organization (WHO), government officials and hardworking scientists worldwide, we can begin to understand some of the details about SARS-CoV-2.

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Investigation of Remdesivir as a Possible Treatment for SARS-2-CoV (2019-nCoV)

Remdesivir (RDV or GS-5734) was used in the treatment of the first case of the SARS-CoV-2 (formerly 2019-nCoV ) in the United States (1). RDV is not an approved drug in any country but has been requested by a number of agencies worldwide to help combat the SARS-CoV-2 virus (2). RDV is an adenine nucleotide monophosphate analog demonstrated to inhibit Ebola virus replication (3). RDV is bioactivated to the triphosphate form within cells and acts as an alternative substrate for the replication-necessary RNA dependent RNA polymerase (RdRp). Incorporation of the analog results in early termination of the primer extension product resulting in the inhibition.

 Note the spikes that adorn the outer surface of the virus, which impart the look of a corona surrounding the virion, when viewed electron microscopically. In this view, the protein particles E, S, M, and HE, also located on the outer surface of the particle, have all been labeled as well. A novel coronavirus virus was identified as the cause of an outbreak of respiratory illness first detected in Wuhan, China in 2019.
This illustration, created at the Centers for Disease Control and Prevention (CDC), reveals ultrastructural morphology exhibited by coronaviruses. Photo Credit: Alissa Eckert, MS; Dan Higgins, MAM CDC

Why all the interest in RDV as a treatment for SARS-CoV-2 ? Much of the interest in RDV is due to a series of studies performed by collaborating groups at the University of North Carolina Chapel Hill (Ralph S. Baric’s lab) and Vanderbilit University Medical Center (Mark R. Denison’s lab) in collaboration with Gilead Sciences. 

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Top 10 Tips to Improve Your qPCR or RT-qPCR Assays

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Scientists have had a love-hate relationship with PCR amplification for decades. Real-time or quantitative PCR (qPCR) can be an amazingly powerful tool, but just like traditional PCR, it can be quite frustrating. There are several parameters that can influence the success of your PCR assay. We’ve highlighted ten things to consider when trying to improve your qPCR results.

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A Molecular Approach to Estimating Time of Death

Stopwatch
I will admit that over the years, I have watched various crime scene investigation shows and read several books by Kathy Reichs and Patricia Cornwell because I was fascinated by forensic science. These same books and shows are a guilty pleasure because as a scientist, I know these portrayals do not accurately reflect how laboratory work is done. Answers are not so cut and dried as an exact estimation of time of death—for example, death was five hours before the body was found in an abandoned warehouse. However, scientists are always looking for ways to improve accuracy in time of death estimates, which are currently based on a few physical clues that are affected by environment and other factors. One approach taken by Sampaio-Silva et al. (1) was to assess the RNA degradation using reverse transcription quantitative PCR (RT-qPCR).

The authors of this PLOS ONE article wanted to determine if nucleic acid degradation could be used as a method to improve time of death estimates. Continue reading “A Molecular Approach to Estimating Time of Death”

Choosing the Right Reverse Transcriptase for Your Project

There are a lot of choices when it comes to reverse transcriptases.  Choosing the correct one for your cDNA synthesis and RT-PCR project is important.    Here are a few questions that will lead you to right RT for your application: Continue reading “Choosing the Right Reverse Transcriptase for Your Project”