Just What Is an RLU (Relative Light Unit)?

Posted on October 28, 2020October 28, 2020 by Promega

This post was contributed by guest blogger, Scott Messenger, Technical Support Scientist 2 at Promega Corporation.

It’s always an exciting time in the lab when you find a new assay to answer an important research question. Once you get your hands on the assay, it is always good to confirm it will work for your experimental setup. Repeating the control experiment shown in the technical manual is a great way to test the assay in your hands.

After running that first experiment of your assay, it looks pretty good. The trends of control and treatment are consistent. Time to get on with the experiments…but wait—the RLUs (Relative Light Units) are two orders of magnitude lower than the example data! I can’t show this data to my colleagues; it doesn’t match. What did I do wrong?

This is a concern that we in Technical Services hear frequently. The concern is real, and I had this same thought when doing some of my first experiments using luminescence. When a question like this comes in, a Technical Service Scientist will make sure the experiment was performed as we described, and in most cases it is. We then start talking about RLUs (Relative Light Units).

Maximize Your Time in the Lab: Improve Experimental Reproducibility with Thaw-and-Use Cells

Posted on October 2, 2020October 7, 2020 by Michele Arduengo

Many cell biology researchers can name their department’s or institutions’s “cell culture wizard”—the technician with 20+ years of experience whose cell cultures are always free from contamination, exhibit reliable doubling rates and show no phenotype or genotype weirdness. Cell culture takes skill and experience. Primary cell culture can be even more difficult still, and many research and pharmaceutical applications require primary cells.

Yet, among the many causes of failure to replicate study results, variability in cell culture stands out (1). Add to the normal challenges of cell culture a pandemic that shut down cell culture facilities and still limits when and how often researchers can monitor their cell culture lines, and the problem of cell culture variability is magnified further.

Treating Cells as Reagents

A good way to reduce variability in cell-based studies is to use the thaw-and-use frozen stock approach. This involves freezing a large batch of “stock” cells, then performing quality control tests to ensure they respond appropriately to treatment. Then whenever you need to perform an assay, just thaw another vial of cells from that batch and begin your assay—just like an assay reagent! This approach eliminates the need to grow your cells to a specific stage, which could take days and introduce more variability.

Tips for Attendees: Making the Most of a Virtual Conference

Posted on July 31, 2020July 30, 2020 by Promega

Today’s blog was written by guest bloggers Tara Luther, Marketing Specialist Genetic Identity, and Allison Suchon, Manager of Tradeshows and Events at Promega.

2020 has been a year of changes for all of us. We’ve learned how to keep in touch while physically distancing. We’ve learned how to work from home with furry coworkers who encourage us to break from the traditional 9–5 routine. We’ve learned how to make changes to our labs to stay safe and productive.

For many of us, this will also be the first time that we attend a virtual conference. While it’s easy to focus on what we’ll be missing by not gathering together, there are advantages to moving to the virtual space. By making the most out of your virtual experience, you’ll be able to walk away with valuable insights, a robust network, and insights that you can use in your own lab.

To help, we’ve put together a list of tips that will help you maximize your experience at any virtual conferences you attend.

Getting Back to the Bench

Posted on July 6, 2020July 6, 2020 by Promega

Today’s blog is written by Technical Services Scientist, Joliene Lindholm, PhD.

Many of us have come back to the lab after a summer of field work or a vacation break, but there is usually someone checking in on the lab to make sure the gel electrophoresis box did not completely overflow with dead bugs and the water baths are not completely overrun with exciting new algae. Maybe this was just because I worked in an older building in an entomology department, but why do insects like running buffer so much? Some labs have been completely shut down for months at this point or maybe just a few essential people have been in keeping stocks and colonies going. Some labs have adapted to the new normal and developed guidelines to keep researchers safe while still doing essential work in the lab. See how the Promega Scientific Applications group has maintained this balance.

Headed back to the lab bench? Take some time to make sure you have everything you need to start up your research projects.

Here are a few tips from what I learned in managing a lab after a period of field work to get back into the swing of things:

Getting a PhD in Sweatpants: Guest Blog by Dr. Susanna Harris

Posted on April 28, 2020April 28, 2020 by Promega

Today’s blog is guest-written by Susanna Harris, who recently defended her PhD thesis at the University of North Carolina in Chapel Hill.

I just defended my PhD. Nearly six years of blood, sweat, and tears, most of which were cleaned up with Kimwipes while sitting at my desk in a laboratory facing out towards the UNC Chapel Hill football field. Nearly six years of work, all summed up in a handful of slides. Nearly six years of work, explained to my friends, family, and colleagues – a moment I had dreamed of since the fall of 2014.

What I hadn’t dreamed of? That I would be sitting at my small desk in the corner of my room, with no present audience aside from my snoring dogs. That there would be no dinner celebration that carried into a night of fun along Franklin Street. That, unseen by the viewers of my defense, I would be wearing sweatpants as my name changed from Ms. to Dr. Harris.

Pictured: The audience for Susanna’s thesis defense.

Why did I wear sweatpants when I could have worn literally anything in my closet? Because I think it’s hilarious. I believe this situation will end and we will walk away with memories and lessons learned from an extremely difficult time in the history of the world. I want to walk away with one more ridiculous story to add to a long list of “What even was that?” tales from grad school.

Working towards a PhD is hard at any time; let’s not pretend this pandemic isn’t making things even worse. I was fortunate in many ways that my advisor had already moved our laboratory to a new state in 2019, allowing me to adjust to meeting through webcams and working from home before the pandemic changed the lives of all North Carolinians. This has given me a unique perspective to tease out which problems come from distance working and which are the result of Safer-At-Home orders. Based on my experiences, here are a few tips, tricks, and words of warning.

Improving Science Literacy for the New Decade

Posted on December 30, 2019December 30, 2019 by Kari Kenefick

Science touches our lives, daily. But far too many scientific concepts and terms are misunderstood and used incorrectly. Even those of us wearing a “scientist” badge sometimes misappropriate terms, which can act to reproduce the misuse.

A basic level of science literacy is so important for all of us. Why? So that when bombarded with comments about vaccination or climate change on a social media site, we are able to sift through the jargon, understand what’s correct and what is not correct, and make decision based in facts vs. internet gossip. With just a bit of knowledge of basic science terms, you are better protected against deception and you’ll know how to sort facts from fiction.

Here are a few general science terms that are commonly misunderstood and misused.

Cloning Blunt-Ended DNA Fragments is Hard: pGEM®-T Vectors Can Make It Easier.

Posted on December 13, 2019January 29, 2020 by Leah Cronan

PCR amplification with a proofreading polymerase, like Pfu DNA polymerase, will leave you with a blunt end. However, another thermostable DNA polymerase, like Taq DNA Polymerase, adds a single nucleotide base to the 3’ end of the DNA fragment, usually an adenine, creating an “A” overhang. This “A” overhang can create difficulties when cloning the fragment is your end goal. You might consider creating a blunt end with Klenow or adding restriction sites to the ends of your PCR fragment by designing them in your primers. But why go through all those extra steps, when that “A” overhang allows efficient cloning of these fragments into T-Vectors such as the pGEM^®-T Vectors? Fewer steps? Who can argue with that?

Tips for Successful Dual-Reporter Assays

Posted on December 6, 2019December 6, 2019 by Ben Schmidt

Previously, we described some of the advantages of using dual-reporter assays (such as the Dual-Luciferase®, Dual-Glo® Luciferase and the Nano-Glo® Dual-Luciferase® Systems). Another post describes how to choose the best dual-reporter assay for your experiments. For an overview of luciferase-based reporter gene assays, see this short video:

These assays are relatively easy to understand in principle. Use a primary and secondary reporter vector transiently transfected into your favorite mammalian cell line. The primary reporter is commonly used as a marker for a gene, promoter, or response element of interest. The secondary reporter drives a steady level of expression of a different marker. We can use that second marker to normalize the changes in expression of the primary under the assumption that the secondary marker is unaffected by what is being experimentally manipulated.

While there are many advantages to dual-reporter assays, they require careful planning to avoid common pitfalls. Here’s what you can do to avoid repeating some of the common mistakes we see with new users:

Continue reading “Tips for Successful Dual-Reporter Assays”

T-Vector Cloning: Questions, Answers and Tips

Posted on November 12, 2019November 12, 2019 by Leah Cronan

Blue/White colony screening helps you pick only the colonies that have your insert.

Q: Can PCR products generated with GoTaq DNA Polymerase be used to for T- vector cloning?

A: Yes. GoTaq® DNA Polymerase is a robust formulation of unmodified Taq Polymerase. GoTaq® DNA Polymerase lacks 3’ →5’ exonuclease activity and displays terminal transferase activity that adds a 3′ deoxyadenosine (dA) to product ends. As a result, PCR products amplified using GoTaq® DNA Polymerases (including the GoTaq® Flexi and GoTaq® G2 polymerases) will contain A-overhangs which makes them suitable for T-vector cloning with the pGEM®-T (Cat.# A3600), pGEM®-T Easy (Cat.# A1360) and pTARGET™ (Cat.# A1410) Vectors.

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Just What Is an RLU (Relative Light Unit)?

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Maximize Your Time in the Lab: Improve Experimental Reproducibility with Thaw-and-Use Cells

Treating Cells as Reagents

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Tips for Attendees: Making the Most of a Virtual Conference

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Getting Back to the Bench

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Getting a PhD in Sweatpants: Guest Blog by Dr. Susanna Harris

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Improving Science Literacy for the New Decade

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Cloning Blunt-Ended DNA Fragments is Hard: pGEM®-T Vectors Can Make It Easier.

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Tips for Successful Dual-Reporter Assays

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T-Vector Cloning: Questions, Answers and Tips

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Treating Cells as Reagents

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