Author: Kelly Grooms

When DNA Is Not Enough: New Research Suggests Epigenetic Factors Play an Important Role in the High Mortality Rate of the Devi Facial Tumor Disease

Posted on by Kelly Grooms

tazThe Devil Facial Tumor Disease (DFTD) is a contagious cancer in Tasmanian Devils that is threatening the species with extinction. This disease is spread from individual to individual and has a 100% mortality rate. It is so deadly because, although the DFTF cells should be attached and killed by the host devil’s immune system, for some reason they are not—and no one is sure why. A study published in PNAS in March of last year (1) showed that DFTD cells don’t express surface MHC molecules. MHC class I and class II molecules are crucial for proper immune response, and their absence on the cell surface could explain why the DFTD cells do not stimulate an immune response.

The authors found that the loss of MHC expression is maintained as the cells divide, and is not a result of structural mutations in the genes responsible for MHC expression. Instead the authors found that this down regulation was the result of regulatory changes including epigenetic modifications to histones. Continue reading “When DNA Is Not Enough: New Research Suggests Epigenetic Factors Play an Important Role in the High Mortality Rate of the Devi Facial Tumor Disease”

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.

RNAi: The Dream Makes a Comeback

Posted on by Kelly Grooms
This Promega Notes Cover from 2004 celebrated the potential stop and go power of DNA-directed RNAi.
This Promega Notes Cover from 2004 celebrated the potential stop and go power of DNA-directed RNAi.

In the early 2000s, RNAi was a hot topic. The science world was abuzz with all the possibilities that harnessing this natural process could hold. And why not? The idea of posttranscriptionally silencing genes using only a small fragment of double-stranded RNA is huge—big enough to earn the scientists who discovered it a Nobel Prize in 2006.

The process of RNAi starts with short (~70 nucleotieds), double-stranded fragments of RNA called short hairpin RNAs (shRNA). These shRNAs are exported into the cytoplasm and cleaved by the enzyme Dicer into smaller pieces of RNA that are about 21 nucleotides long and are referred to as small interfering RNAs (siRNA). The siRNAs reduce or stop expression of proteins through a sequence of events where the antisense strand of the siRNA is incorporated into and RNA-induced silencing complex (RISC), which then attaches to and degrades its complimentary messenger RNA, thereby reducing or completely stopping expression.

It turned out, however, that harnessing the promise of RNAi was a little trickier than anticipated. Continue reading “RNAi: The Dream Makes a Comeback”

Chikungunya Virus and the Promise of a Virus-Like Particle Vaccine

Posted on by Kelly Grooms

My family and I just returned from a week-long camping trip along the North Shore of Lake Superior in Minnesota. It is beautiful country, filled with lakes, rivers, ponds—and mosquitoes, lots and lots of mosquitoes. We went prepared for the worse. We had a screen tent, head nets and tubes and tubes of insect repellent because in this area of the world, mosquitoes are a flying, buzzing, picnic-ruining, itch-inducing pest. In the US, though, a pest is really all they are. In other areas of the world they are a flying, buzzing, disease-carrying, deadly menace.

Image courtesy of James Gathany and the CDC
Image courtesy of James Gathany and the CDC

Mosquitos act as vectors for many diseases including malaria, Dengue fever, Yellow fever, encephalitis, West Nile Virus and chikungunya virus. Many of these diseases are deadly; in fact, mosquitoes are responsible for more human deaths than any other animal (~725,000 deaths annually). Although most of these diseases have a long and infamous history, two of them, West Nile virus (first identified in 1932) and chikungunya virus (first identified in 1950), are relative new comers on the world health stage. Continue reading “Chikungunya Virus and the Promise of a Virus-Like Particle Vaccine”

Discovering the Complexity of the Human Proteome

Posted on by Kelly Grooms

Transcription TranslationI should preface this blog by stating that I am a nucleic acids gal. My years in the lab were spent with tubes of DNA and RNA. In fact my one and only tentative foray into the field of protein resulted in a Western Blot so ugly that those who witnessed it have been sworn to secrecy. Given all of this, the mapping of the human proteome might seem like an odd topic for me to write about. Except that it isn’t really, because the sequencing of the proteome offers answers to some of the questions that the sequencing of the genome didn’t.

First, let’s start with what a proteome is: A proteome is all the proteins expressed at a certain time point. It can be as limited as the proteome of a single cell or as all encompassing as the proteome of an entire genome. However, unlike the genome, which is genetic information encoded in an organism’s DNA or RNA, the makeup of a proteome can vary dramatically as a result of expression patterns, alternative splicing events and post-translational modifications.

The genome is a constant, what you see today is what will still be there tomorrow. The proteome, on the other hand, is a constantly changing landscape. Up regulation or down regulation of a gene can mean more or less protein is present. Alternative splicing and post-translational modifications can result in fundamental changes to the protein itself.

In other words, if the genome is a beautiful, pristine Ansel Adams print, then the proteome is that same scene as interpreted by Andy Warhol—in Technicolor and 3D. Continue reading “Discovering the Complexity of the Human Proteome”

From Soot, Turpentine and Walnut Oil to Bio-Inks with Living Cells: Printing Reinvents Itself in 3D and Comes Alive

Posted on by Kelly Grooms
Gutenberg-Style Printing Press.
Gutenberg-Style Printing Press (Replica). Wikimedia Commons.

Printing has been an integral part of society since the first movable type printing press was invented by Johannes Gutenberg in the 15th century. And yet, many recent headlines have heralded its impending death.  As everything from books to newspapers transitioned to digital media, the traditional printing techniques seemed destined for the history books and museums. However, even before the popularity of printing started to fade, some of the concepts were being reimagined in amazing, new—and three dimensional— ways. Continue reading “From Soot, Turpentine and Walnut Oil to Bio-Inks with Living Cells: Printing Reinvents Itself in 3D and Comes Alive”

Battling Obesity One Thermostat at a Time

Posted on by Kelly Grooms

ThermometerWinter in Wisconsin is synonymous with cold, and this year thanks to the “wobbly polar vortex” it has been really, really cold. I have been very grateful for my under-desk space heater at work and my toasty gas fireplace at home. However, according to an article currently in press in the journal Trends in Endocrinology and Metabolism (1), all that lovely toasty warmness might be working against me if I am hoping to loose weight.

Excess weight is nothing more than a positive energy balance, meaning we have taken in more calories than we have burned. The deceptively simple sounding solution for losing or maintaining weight is to take in no more calories than you will be expending. Typically this is achieved by eating less, increasing physical activity or through pharmacological interventions. However, anyone who has ever tried any of these approaches knows that there is nothing simple about them, and often times the results are disappointing or short lived.

The authors of the Trends in Endocrinology and Metabolism paper hypothesize that regular exposure to mild cold could affect our energy expenditure in a positive way. We know that shivering produces heat (shivering thermogenesis) and thus expends energy. It can increase the metabolic rate to up to five  times the resting rate (2). However, it is not comfortable and can make coordinated movements difficult. The authors focused on something close to this state, but not quite. Nonshivering thermogenesis (NST) is the cold-induced increase in heat production that is does not involve muscle activity like shivering. It occurs by activating brown adipose tissue (3–5).

Although there is great variation between individuals, most young and middle-aged people will see an increase in NST between a few percent to up to 30% when exposed to mildly cold conditions (1). The authors note that a recent study found a significant decrease in body fat content following a 6 week cold acclimation study (2 hours/day at 62.6°F [17°C]; 6).

ScaleAs many of us can probably attest, the downfall of undertaking anything that increases our body’s energy usage is that often we end up increasing our caloric intake to compensate for it. Interestingly, the authors point to a 2009 study (7) that found evidence that increased food intake did not fully compensate for this type of cold-induced thermogenesis.

Personally, I love to be warm— as evidenced by my previous comments about space heaters and gas fireplaces. However, if the benefits of cold acclimation suggested by this paper hold to be true, I think I could tolerate 2 hours a day at 62°C.

References

  1. Van Marken Lichtenbelt. W. et al. (2014) Cold exposure—An approach to increasing energy expenditure in humans. Trends Endrochron. Met. In Press.
  2. Jansky, L. (1998) Shivering. In Physiology and Pathophysiology of Temperature Regulation (Blattheis, C.M. ed.) World Sceintific.
  3. Cannon, B. and Nedergaard, J. (2004) Brown adipose tissue: Function and physiological significance. Physiol. Rev. 84, 277–359.
  4. Van Marken Lichtenbelt, W.D. et al. (2009) Cold-activated brown adipose tissue in health adult men. N. Engl. J. Med. 360, 1500–1508.
  5. Virtanen, K.A. et al. (2009) Functional brown adipose tissue in health adults. N. Engl. J. Med. 360, 1518–1525.
  6. Yoneshiro, T.  et al. (2013) Recruited brown adipose tissue as an antiobesity agent in humans. J. Clin. Invest. 123, 3404–3408.
  7. Cannon, B. and Nedergaard, J. (2009) Thermogenesis challenges the adipostat hypothesis for body-weight control. Proc. Nutr. Soc. 68, 401–407.

Cell-free Expression: A System for Every Need

Posted on by Kelly Grooms

6634MA

Cell-free protein expression is a simplified and accelerated avenue for the transcription and/or translation of a specific protein in a quasi cell environment. An alternative to slower, more cumbersome cell-based methods, cell-free protein expression methods are simple and fast and can overcome toxicity and solubility issues sometimes experienced in traditional E. coli expression systems. Continue reading “Cell-free Expression: A System for Every Need”

Inheriting Fear: Mice Haunted by Parent’s Fears

Posted on by Kelly Grooms

baby miceImagine that you are sitting in your room when you smell cherries and you are suddenly, inexplicably afraid. Although odors can elicit strong emotional responses, you have no bad memories of cherries. What you don’t know is that your father did, and you have inherited his fear. Sound far fetched? Maybe not. A paper published in Nature Neuroscience found just such an inherited association in mice (1). Continue reading “Inheriting Fear: Mice Haunted by Parent’s Fears”