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:
This past weekend was the 9th Annual Wisconsin Science Festival, and we at Promega were excited to join in the celebration of science throughout the state. We participated in the Discovery Expo on Thursday and Friday, where dozens of demonstrations and exhibits were scattered throughout the Wisconsin Institute for Discovery building. Thousands of children on field trips filled the halls, eager to poke and prod at strange and exciting new things.
At our table, we talked about the science of bioluminescence. With 3D-printed firefly luciferase models in hand, we showed the glow of recombinant luciferase to the incoming children and explained to them how scientists could use bioluminescence like a tiny “flashlight” to look inside of cells and watch what’s happening. Our learners received a nice little reward for their attentiveness in the form of glow-in-the-dark firefly stickers.
The stage is set. You’ve spent days setting up this experiment. Your bench is spotless. All the materials you need to finally collect data are laid neatly before you. You fetch your cells from the incubator, add your detection reagents, and carefully slide the assay plate into the luminometer. It whirs and buzzes, and data begin to appear on the computer screen. But wait!
Don’t let this dramatic person be you. Here are 8 tips from us on things to watch out for before you start your next luminescent assay. Make sure you’ll be getting good data before wasting precious sample!
Tailing blunt-ended DNA fragments with TaqDNA Polymerase allows efficient cloning of these fragments into T-Vectors such as the pGEM®-T Vectors. This method also eliminates some of the requirements of conventional blunt-end cloning — Fewer steps, who can argue with that?
We spend a lot of time looking at history and imagining—”what was it like when…?” As a biologist, I find myself most drawn to stories about the evolution of life. Why does this plant have purplish leaves? How did this species end up in a symbiotic relationship with this other species? How did this animal get to this tiny island 20 miles off the Southern coast of Iceland?
That last one was too specific to be rhetorical, wasn’t it? The volcanic island of Surtsey broke the ocean surface on November 14, 1963, and continued to erupt until June 5, 1967, reaching its maximum size of 2.7 km2 (about the size of Central Park in New York City). At this size, it was large enough to be a good site for biocolonization. Only a few scientists are allowed to visit the island, ensuring that colonization of the island can occur without human interference. Continue reading “Science Visitors Only: Watching Life Grow on a New Island”
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.
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!
We have published 130 blogs here at Promega this year (not including this one). I diligently reviewed every single one and compiled a list of the best 8.5%, then asked my coworkers to vote on the top 5 out of that subset. Here are their picks:
This past weekend, I had the opportunity to be a part of “Once Upon a Christmas Cheery in the Lab of Shakhashiri”. Bassam Z. Shakhashiri is a professor of chemistry at the University of Wisconsin–Madison who is well-known for his fun science demonstrations and a fervent dedication to public science communication. Once Upon a Christmas Cheery started in 1970 as an end-of-semester treat for Dr. Shakhashiri’s freshman chemistry class; by 1973, the Christmas lecture had become so popular that Wisconsin Public Television offered to broadcast it during Christmas week, and this collaboration has continued uninterrupted ever since.
That’s 49 years of Christmas lectures, commemorated by making indium, the 49th element, the Sesame Street-esque “sponsor” of the show. It helps that indium burns bright violet, the name of Dr. Shakhashiri’s granddaughter and hence his favorite color. The color purple made a firm foundation for many aspects of the show: The chrysanthemums frozen in liquid nitrogen were purple, as was the balloon I inflated during my spiel on air movement. Most of the set was various shades of purple, too.
Happy graduation! Whether you graduated last week or twenty years ago, the experience is roughly the same. As soon as you arrive on the far side of the stage, empty diploma folder under your arm, hand still sticky from the Dean’s sweaty handshake, the reality of post-academic life sets in. Perhaps grad school is on the horizon for some and others might be busy prepping for med school. For some of us, though, our years of formal education end after four and we run off to rejoice in our newfound freedom. No more exams, group projects, late nights writing papers, disapproving professors, supervisors and mentors – done with that life forever! We didn’t even bother with the GRE, MCAT, LSAT or a single “Why [insert school]” essay. Now it’s off to enjoy the Real World, which will definitely be better than college.
I’ve found, in my one year of post-college life, that sometimes you can miss academic life. You’ll occasionally look back and think, “I didn’t know how good I had it.” In particular, those of us with a pure love of learning can find ourselves unsatisfied with our prospective learning opportunities or lack thereof. We spent college soaking up mountains of knowledge–and not just from textbooks. University life gives you access to free talks from eminent thought leaders, unrestricted access to myriad scientific journals, and plenty of people around who are eager to argue about that day’s lecture in Cell Biology or Neuroscience. After college, it’s tough to fill that void.
I work at Promega (obviously), a biotech company, so I still have access to journals and there are plenty of brilliant scientists around me. However, I’m still looking for more opportunities to learn and grow. I may be out of school, but the love of science never goes away. Here are a few of my tips for everyone receiving their hard-earned science degree this spring.
The other night I was playing volleyball and, during a team huddle, made a joke that the only players working hard were those with two X chromosomes (a playful jab at the male players on my team). The only response I got was a single, delayed smile along with a bunch of blank looks. That joke certainly would have produced a better reaction among my scientific colleagues, even if that simply meant a bunch of immediate groans.
I happen to think science-minded folks like myself have a terrific sense of humor, it’s just tailored to a more niche audience since a lot of the jokes we tell may not be immediately understood by the average person. While I appreciate comedy in all forms, I delight in laughing at and making jokes related to science.
Since I don’t think I am alone, I thought I would share a few events in today’s blog that really highlight the humor that can be found in the scientific community.