What Things Are You Thankful for in Science?

Posted on by Michele Arduengo, PhD

What are you thankful for in science?
What are you thankful for in science?
As the social media lead for Promega, I keep my eye on trends in new media. I have personal accounts that I keep mostly to see what other people are doing. I try hangouts, social networking and other things so that I have an idea of developing practices outside of the biotechnology industry. One activity that has been popular over the last couple of years during the month of November in the United States is the Facebook post of “30 days of thanksgiving”.

I wondered what “thanksgiving” looks like to the research scientist. So I asked:

What are the things you are thankful for in science?

The answers have been as varied as the people I talked to ranging from little things like water bath floats to really big things, like the renewal of your research funding or achieving tenure.

Here are some of the answers from my informal inquiries:

“Tube floaties for water baths.”

—E.V., genomics product manager

“I was always thankful for Geiger counters.”

—K. G., science writer

“Thermal cyclers and Taq Polymerase. As an undergrad I watched someone sit with a timer and move their tubes between water baths at 3 different temperatures, opening tubes and adding polymerase at the end of each cycle. Modern PCR is SOOO much easier.”

—M.M., research scientist

“I am thankful for competent cells. I remember preparing the CaCl2 and doing slow centrifugation. Also thankful for serum-compatible transfection, rapid ligations and online journal access (no longer have to traipse over to the university library to get papers photocopied- uuurrrgggghhh).”

—R.D., technical services scientist

“How about T-vectors for cloning? I was no molecular biologist, but could make a T-vector work.”

—K.K., science writer

“I am thankful for open-access journals and the ability to read the full article without an institutional subscription.”

—S.K., science writer

“I am ever so thankful for ONLINE ORDERING! So awesome. Throw in online technical manuals, on-line support tools, on-line calculators – all are awesome!!”

—A.P., director, scientific courses

“I am thankful for automated sequencing- manual sequencing was laborious and hazardous!!!”

—R.G., technical services scientist

Do any of these resonate with you? What are you thankful for as a scientist? Let us know in the comments.

DNA Amplification Using Body Heat, No Instrument Required

Posted on by Sara Klink

Cartoon by Ed Himelblau
Cartoon by Ed Himelblau Copyright Ed Himelblau.
When I was in the lab, there was more than a few times I held tubes in my hand (maybe even under my arm) to make them thaw faster, especially reaction buffers. However, I never considered whether this could be a strategy for actual incubation although humans run at about 37°C and many restriction enzyme reactions proceed most efficiently at 37°C. But research published in PLOS ONE by Crannell, Rohrman and Richards-Kortum took this idea and decided to experiment with the possibility of eliminating an instrument-based DNA amplification. Continue reading “DNA Amplification Using Body Heat, No Instrument Required”

Insights into the Function of P7C3 Compounds in Neuroprotection

Posted on by Kari Kenefick
The multiple Lombardi trophies won by Pittsburgh Steelers. Image used under Wikimedia Creative Commons, and attributed to daveynin.
The multiple Lombardi trophies won by Pittsburgh Steelers. Image used under Wikimedia Creative Commons, and attributed to daveynin.

It is fall and the season for American football. For this football fan, watching the game is a bit less enjoyable than it used to be, as more and more information is available about the serious and permanent brain injuries suffered by football players.

In the introduction to a recent paper in the journal Cell, “P7C3 Neuroprotective Chemicals Function by Activating the Rate-Limiting Enzyme in NAD Salvage”, not a word about American football is mentioned.

However, the paper begins, “No substantive therapeutics are available for the treatment of almost any form of disease entailing nerve death” (1). The authors list a range of neurodegenerative disorders such as Huntington’s, Alzheimers and Parkinson’s diseases, as well as ALS  or Lou Gherig’s disease. They also note that there are currently no effective treatments for trauma to the brain or peripheral nervous system.

The authors note that a chemical treatment that could interfere with nerve cell death would have a “transformative impact in modern medicine”. Continue reading “Insights into the Function of P7C3 Compounds in Neuroprotection”

Optimizing Tryptic Digestions for Phosphoproteomics Analysis

Posted on by Gary Kobs

11296971-DC-CR-KinaseProtein phosphorylation is the most widespread type of post-translational modification. It affects every basic cellular process, including metabolism, growth, division, differentiation, motility, organelle trafficking, membrane transport, muscle contraction, immunity, learning and memory (1,2). Protein kinases catalyse the transfer of the phosphate from ATP to specific amino acids in proteins. In eukaryotes, these are usually Ser, Thr and Tyr residues. Due to the development of specific phosphopeptide enrichment techniques and highly sensitive MS instruments, phosphoproteomics has enabled researchers to gain a comprehensive view on the dynamics of protein phosphorylation and phosphorylation based signaling networks.

Due to its high cleavage specificity, trypsin is the commonly used proteolytic enzyme in MS-based proteomics, cleaving peptides carboxyterminal of the amino acids lysine and arginine. However, various factors such as the tertiary structure of a protein, adjacent basic amino acids or negatively charged residues close to cleavage sites as well as PTMs are known to impair proteolysis.

To gain closer insights into the impact of phosphorylation on tryptic digestion, a recent publication(3) systematically characterized the digestion efficiency of model peptide sequences that are known to be prone to incomplete digestion.

The results indicated that increasing trypsin concentrations up to a trypsin to peptide ratio of 1:10 led to a significant gain (1) in the overall number of phosphorylation sites (up to 9%) and in the intensities of individual phosphopeptides, thereby improving the sensitivity of phosphopeptide quantification.

The effect of organic solvents (ACN, acetonitrile and TFE trifuorethanol was also evaluated). Positive results were noted with TFE when determining the digestion of individual peptides. However TFE interfered with TiO2 phosphopeptide enrichment and therefore was not recommended for use with complex samples.

  1. Engholm-Keller, K and Larsen, M.R. (2013) Technologies and challenges in large scale phosphoroproteomics. Proteomics 13, 910–31.
  2. Beausoleil, S. A. et al. (2010) Tissue-specific atlas of mouse protein phosphorylation and expression. Cell 143, 1174–89.
  3. Dickhut, C. et al. (2014) Impact of Digestion Conditions on phosphoproteomics. J. Proteome Res. 13, 2761–70.

Telepathy and Mind Control: From Science Fiction Movies to Reality?

Posted on by Terri Sundquist

BrainInterfaceThere is something very futuristic, and perhaps scary, about the idea of nonverbally transferring one person’s thoughts to another person, especially for the purpose of controlling or influencing a person’s actions and behaviors. Maybe that’s why telepathy and mind control are favorite topics of many science fiction movies. However, there are times when direct, nonverbal transfer of thoughts would be advantageous, for example when communicating complex concepts or feelings that are difficult to convey. Direct transfer also would circumvent the need to translate information from one language to another. For these reasons, scientists are currently developing technologies to allow such thought transfers. A recent PLOS ONE article describes a simple brain-to-brain interface in humans and shows how this interface can be used to capture a thought generated by one person and communicate that information directly to the brain of a second person and elicit a physical response (1).

Continue reading “Telepathy and Mind Control: From Science Fiction Movies to Reality?”

6 + 1 Ways Dual-Reporter Assays Can Save Your Data

Posted on by Kyle Hooper

Updated 02/12/2021

Dual-Reporter-Assay

Transient transfection is often used to perform reporter assays. We have advocated using a dual-reporter system for decades to normalize the data obtained and gain a clearer understanding of your results.  The experimental reporter should vary with treatment and the control reporter should vary little with treatment. The control reporter thus serves as a marker to help you understand the relative activity of your experimental reporter. The bioluminescent Dual-Luciferase® method allows for sequential detection of the second reporter in a single sample providing a simple two-step normalization method. Here are seven ways in which dual-reporter assays help you avoid misinterpreting results.

Simply comparing the ratio of the experimental to the control reporter can resolve differences in:

  1. Number of Cells/Well: When manually pipeting cells into a 96-well plate, there is always a chance of having variable numbers of cells in each well. This variation is cell number will affect the experimental and control reporters equally, so the ratio of experimental:control reporter activity will eliminate false interpretation of the experimental data–whether it affects an entire row or column on the plate or individual wells.
  2. Transfection Efficiency: The variations in transfection efficiency will equally affect both the experimental and control reporters so the ratio of activity in dual-reporter assays will normalize the data.
  3. Cell Viability: Often, reporter assays look at the dose response curve of a particular compound with regard to gene expression. Ideally, if a compound causes a change in the experimental reporter the control reporter will demonstrate little effect. However, if the compound is toxic, both the experimental and control will be altered and the ratio will tell you whether the compound truly affects reporter activity or just kills the cells.
  4. Lysis Efficiency: When lysing a plate of cells, you could encounter situations where rows or columns lyse differently, especially if you are using manual disruption or get interrupted mid-plate. The difference is lysis will affect the experimental and control equally so the ratio will remove the variation.
  5. Temperature: Ideally, a plate should be equilibrated to ambient room temperature before proceeding to the reporter assay. Plates can cool at different rates or researchers anxious to record data may read the data early. Temperature variations will affect both reporters so the ratio will limit the affect on the data.
  6. Measurement Time: Repetition of data is a hallmark of good science. You are often called upon to repeat experiments sometimes days or weeks apart. Let’s say you repeat your experiment one week after the initial experiment. The first time you measured the response, you waited 10 minutes after reagent addition to read, this week you waited 30 minutes. This will affect both reporters equally and therefore the ratio will allow you to more easily compare the data from this week and last week.

Bonus Benefit from Dual-Luciferase®, Dual-Glo® and the NanoGlo® Dual Luciferase Reporter Systems: No Lysate Splitting: Promega dual-reporter assays are designed for same-well multiplexing so there is no chance of variations creeping into your data due to unequal splitting of the cellular lysate to measure two separate reporter activities.

Since the introduction of the first bioluminescent dual-luciferase assay in 1995, this approach has been used in countless studies to advance our scientific understanding of cellular gene regulation.

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Biotechnology Youth Apprenticeship Program Fosters Young Scientists

Posted on by Barbara Bielec

Student working in laboratory.
Photo credit: BTC Institute.
Ellyn Lepinski is an intern at Promega who started her biotechnology career path five years ago as a high school junior taking a course from the BTC Institute (www.btci.org) as part of the Biotechnology Youth Apprenticeship Program.

Ellyn credits the program with helping her achieve her goals:

“Over the course of two years in which I was a Youth Apprentice, I obtained numerous skills, both inside and outside of the lab. I gained valuable scientific experience, including techniques like gel electrophoresis, nucleic acid purification, PCR, SDS-PAGE, Western blotting, cell culture and more.

On a personal level, I became very close with other students in the class and with our instructors, Barbara Bielec and Chad Zimprich. Everyone involved was always very approachable and willing to help with both laboratory tasks and in terms of giving advice for the future.

Through the program, I was placed in Dr. Que Lan’s entomology lab at UW-Madison, beginning in 2009. While there, I worked on a project involving sterol carrier protein-2, a protein involved in cholesterol uptake in mosquitoes.Notably, I am still working in Dr. Lan’s lab, however my research focus has shifted to bacterial fermentation. In between working in Dr. Lan’s lab, I also worked at the Forest Products Laboratory (USDA).

Additionally, this past June, I began an internship at Promega in the Scientific Applications department. Here I work to develop new applications for existing projects. This November marks five years of laboratory research for me, which would not have been possible without the Youth Apprenticeship Program and everyone involved. In addition to the specific labs that I have had the opportunity to work in, my experience in the Youth Apprenticeship Program has allowed me to emerge as a leader in my college lab courses. The program has clearly made a phenomenal impact on my life and is something I am very grateful for.”

Photo credit: BTC Institute.
Photo credit: BTC Institute.
Since 1993, the BTC Institute in partnership with the Dane County School Consortium has helped make such opportunities possible to nearly 300 students from public schools throughout Dane County. The program includes a paid apprenticeship in an industry or UW-Madison research lab and specialized instruction. In addition to being paid for their work, students receive high school credit for their participation in the worksite and the specialized biotechnology course held at the BTC Institute.

One aspect of the program that makes it so effective and unique is the amount of time that students spend working. Youth apprentices who start as juniors in the program must work 900 paid work hours to earn the Science, Technology, Engineering and Math (STEM) Skill Standards Certificate from the State of Wisconsin, youth apprentices who start work as seniors must earn 450 work hours. Students have had employment at a variety of companies and UW-Madison research labs, a few examples that have hired multiple apprentices include Genus PIC (ABS), MOFA Global, Promega and laboratories in the UW-Madison Departments of Bacteriology, Biochemistry, Entomology, Genetics, Horticulture, Plant Pathology and Surgery. Many of the students, like Ellyn, continue to be employed by their worksite long after they graduate from high school—proof of how effective this program is in helping to create the next generation STEM workforce.

Each year the BTC Institute hosts a Youth Apprenticeship Program preview night for all of the Dane County youth apprenticeship options: biotechnology, automotive technician, health services, and many more (www.dcsc.org). This year the preview nights will be held February 24 and 25 starting at 5:00pm. Students in grades 10 and 11 who are interested in learning more about the program are encouraged to attend one of the evening sessions with a parent.

Genome Editing and Reporter Technologies Enable Endogenous Pathway High-Throughput Assays

Posted on by Promega

ImageSource=RCSB PDB; StructureID=1qpf; DOI=http://dx.doi.org/10.2210/pdb1qpf/pdb;
ImageSource=RCSB PDB; StructureID=1qpf; DOI=http://dx.doi.org/10.2210/pdb1qpf/pdb;


This article review was written by guest author, Amy Landreman, in the Cellular Analysis and Proteomics Group at Promega.

Charcot-Marie Tooth (CMT) disease is one of the most common inherited neurological disorders affecting approximately 2.8 million people worldwide. The most common form of CMT, CMT Type 1A, is caused by a 1.5Mb genomic duplication on Chr17 that results in trisomy of the critical myelin gene Peripheral Myelin Protein 22 (Pmp22). The extra copy of Pmp22 results in excessive PMP22 protein causing the neurophathy associated with CMT type 1A. Although there is no way to remove the extra copy of the gene, even subtle decreases in Pmp22 expression have shown promise against this inherited neuropathy in laboratory models.

In a recent paper, Inglese et al. 2014, describe an interesting new approach used to identify compounds that effectively decrease Pmp22 expression using a novel gene editing strategy and reporter-based screen. Their challenge was to create an assay that accurately represented endogenous Pmp22 expression including both transcriptional and post-transcriptional regulatory mechanisms, while maintaining the sensitivity required to detect subtle changes in expression in a loss of signal assay in a format compatible with microtiter 1536-well quantitative high-throughput screening (qHTS). Continue reading “Genome Editing and Reporter Technologies Enable Endogenous Pathway High-Throughput Assays”

Tips for a Social Media User

Posted on by Sarah Dewbre

Social%20Networks_smallerAs an HR professional, I attend different seminars and conferences to obtain credits for my HR certification. We had a SHRM (Society for Human Resources Management) state conference last week, and I learned all kinds of new strategies involving recruitment, succession planning, employee engagement and change management. One topic was present in every session I was in: social media. How to recruit through social media, engage employees in social media, and how to maintain your company and personal brand through social media.

One session in particular was focused solely on social media, and how it affects the processes of hiring, discipline and discharge. Continue reading “Tips for a Social Media User”

Choosing Your Subcloning Strategy

Posted on by Kelly Grooms

Before you begin your subcloning, you need to know: The restriction enzyme (RE) sites available for subcloning in your parent vector multiple cloning region (or in the insert if you need to digest the insert); the RE sites available in the destination vector multiple cloning region (MCR); and if these same sites also occur in your insert. Once you know this information, you can use the chart below to decide which subcloning strategy to use.

4498MA-[Converted]

To learn more about subcloning, visit our Subcloning Notebook.