Promega is a chemistry and instrument supplier to scientists in diverse industries and research labs around the world. True. But we are more than just a supply company; we are scientists dedicated to supporting the work of other scientists. We want the science behind the technologies we develop to be both vetted and valued by the scientific community at large, which is one reason our scientists take the time to prepare and submit manuscripts to peer-reviewed journals. Here we call out some of our published research papers that were highly read in 2019. In the journal ACS Chemical Biology alone, five Promega-authored papers were among the top 10 most read papers in 2019. Here’s a quick review of the highlights from these ACS papers.
Understanding the expression, function and dynamics of
proteins in their native environment is a fundamental goal that’s common to
diverse aspects of molecular and cell biology. To study a protein, it must
first be labeled—either directly or indirectly—with a “tag” that allows
specific and sensitive detection.
Using a labeled antibody to the protein of interest is a
common method to study native proteins. However, antibody-based assays, such as
ELISAs and Western blots, are not suitable for use in live cells. These
techniques are also limited by throughput and sensitivity. Further, suitable
antibodies may not be available for the target protein of interest.
As the number of children diagnosed with autism spectrum disorder (ASD) continues to rise, the search for a cause continues. Scientists have been studying genetically modified oxytocin receptors, which have shown promise as a target for studying ASD-related behaviors. One of the obstacles to designing robust scientific experiments for investigating potential ASD causes or treatments is the lack of a truly appropriate model organism for social behaviors in humans (1). Sure, there are the traditional lab rats and lab mice that demonstrate a certain level of social behaviors. However, there has been a loss of natural social behaviors in common lab mice strains because of the reduction in genetic complexity from inbreeding and adaptation to captivity (2). These animals cannot fully represent the depth of human social behaviors, including the ability of humans to form lasting social bonds (1).
Last week, a diverse group of stakeholders attended CRISPRcon Midwest, hosted by the Keystone Policy Center and the University of Wisconsin–Madison. The goal of the day-long conference was to emphasize the importance and value of gene editing technology, and how it must be communicated deliberately between scientists, the public, policymakers, and other stakeholders.
Julie Shapiro, Senior Policy Director of Keystone Policy Center, acted as Emcee for the event. Given the diverse group of attendees, she mentioned in her opening remarks that the event organizers were “seeking conversation, not consensus” and emphasized the “power of respectful dialogue.” A slide overhead showcased the ground rules for the day, which included statements such as “dare to listen, dare to share, and dare to disagree.”
CRISPRcon aimed to included voices beyond those represented by keynote speakers and panelists, so they incorporated live polling through an online app to keep the audience engaged and an active participant in the conversations throughout the day. From the opening remarks, it was clear that this conference would not just deliver on its promise of thoughtful conversation about the science, but build further understanding about the societal impacts of a rapidly advancing technology.
With the advent of genome editing using CRISPR-Cas9, researchers have been excited by the possibilities of precisely placed edits in cellular DNA. Any double-stranded break in DNA, like that induced by CRISPR-Cas9, is repaired by one of two pathways: Non-homologous end joining (NHEJ) or homology-directed repair (HDR). Using the NHEJ pathway results in short insertions or deletions (indels) at the break site, so the HDR pathway is preferred. However, the low efficiency of HDR recombination to insert exogenous sequences into the genome hampers its use. There have been many attempts at boosting HDR frequency, but the methods compromise cell growth and behave differently when used with various cell types and gene targets. The strategy employed by the authors of an article in Communications Biology tethered the DNA donor template to Cas9 complexed with the ribonucleoprotein and guide RNA, increasing the local concentration of the donor template at the break site and enhancing homology-directed repair. Continue reading “All You Need is a Tether: Improving Repair Efficiency for CRISPR-Cas9 Gene Editing”
G protein-coupled receptors (GPCRs) are a large family of receptors that traverse the cell membrane seven times. Functionally, GPCRs are extremely diverse, yet they contain highly conserved structural regions. GPCRs respond to a variety of signals, from small molecules to peptides and large proteins. Many GPCRs are involved in disease pathways and, not surprisingly, they present attractive targets for both small-molecule and biologic drugs.
In response to a signal, GPCRs undergo a conformational change, triggering an interaction with a G protein—a specialized protein that binds GDP in its inactive state or GTP when activated. Typically, the GPCR exchanges the G protein-bound GDP molecule for a GTP molecule, causing the activated G protein to dissociate into two subunits that remain anchored to the cell membrane. These subunits relay the signal to various other proteins that interact with or produce second-messenger molecules. Activation of a single G protein can result, ultimately, in the generation of thousands of second messengers.
Redundancy equips us to survive. We have more than one lung or one kidney for a reason—if one organ in a pair gets damaged, we can still manage if the other is functional. At the cellular level, we have two copies of each chromosome in every non-germline cell. Each copy was inherited originally from a single sperm and ovum, which are “haploid” cells. Consequently, there are two copies of any given gene in non-germline “diploid” cells. In many cases, should one copy of a gene be damaged, the cell can still survive with the other, functional copy of a gene. In plants, this redundancy is common, and many plants exhibit polyploidy. In an extreme example of polyploidy, the large (by bacterial standards) but otherwise unassuming species Epulopiscium contains tens of thousands of copies of its genome.
Getting DNA or RNA into cells can be a tricky business, and a variety of transfection reagents have been developed over the years to make the process easier. Lipid-based reagents are especially popular because they combine efficient transfection with relatively low toxicity.