...the nucleus, also called the 4D nucleome. Visualizing specific loci has historically been difficult, as...successfully resolved loci with physical map distances of 2 and 75 Mbp, with the calculated fluorescent distances...to label certain loci, but this approach is not scalable for every locus...unlike CRISPR. Early CRISPR ...Figure 1: CRISPRainbow was used to track the localization of 6 chromosome-specific loci in the U20S cancer... every genomic locus, and to do so in vivo. Catalytically dead Cas9 (dCas9) cannot induce gene editing... to a given genomic locus, you can track the localization and movement of this locus in live cells! This...distances correlating with the previously established physical map. In comparing pairs of targets ~2 Mbp apart...

... brainstorming new technological innovations. Let’s take a look at three historical techniques: southern...computers, cell phones, and cars become more technologically advanced leaving earlier versions obsolete,...that save both time and money. Yet, knowledge of historical techniques comes in handy whether you are perusing...cloning or knockout experiments to forensics and clinical diagnostics. One example of southern blotting ...fragments of a DNA sequence of interest to create a physical map of the DNA. Starting in the 1970s, restriction...keep restriction enzymes on hand for cloning and analytical digests. A researcher can test the orientation...). Sanger sequencing uses modified nucleotides, called dideoxynucleotides (ddNTPs), lacking the 3’ hydroxyl...

...Sensitivity in Western Blotting. Journal of Pharmacological and Toxicological Methods, 64, 233–237. https://doi....to get right. If you’ve thoughtfully done the technical design of your blot, but are still finding yourself... be done at 4 °C or on ice, whichever is more practical. If your lysing method is not resulting in your...Most people will simply load an extra ladder or technical replicate (ie, leftover sample) to fill the relevant...you may want to consult Tables 1 and 2 in our Technical Design of a Western Blot post to make sure you...transfer. Troubleshooting The two most common (non-technical) issues with transferring are smaller proteins...than just high background, your issue is likely technical. White circles, for example, could indicate that...

...UCIBIO) focused on understanding the physiological and pathological roles of human cell glycans. Accordingly... biosynthetic pathway and have a wide range of clinical manifestations. PMM2-CDG is the most frequent ...reported that PMM2-CDG patients suffer from immunological defects. These are characterized by a predisposition...these lines to study how immune processes and biological pathways are altered in PMM2-CDG. These studies...reducing their risks for infection and other pathological conditions. References: Monticelli, M., Ferro... J., dos Reis Ferreira, V. & Videira, P. A. Immunological aspects of congenital disorders of glycosylation...Biology at the University of Colorado Anschutz Medical Campus in Aurora, Colorado, USA. The primary focus...

...notable in recent years. A variant of V-U, paradoxically called V-K, is a CRISPR-associated transposase ... CRISPR-Cas systems. Philosophical Transactions of the Royal Society B Biological Sciences, 374(1772),...What’s in a type? That which we call CRISPR, by any other name would…probably still edit genomes. Understanding...function and composition; subtypes and variants are typically based on species of origin. There are currently...systems. Type III systems are unique, as they can technically target both RNA and DNA, though it is largely...systems function, a III-E effector was developed, called Cas7-11. Despite being in Class 1, Cas7-11 is a... that other CRISPR types do, due to a lack of canonical CRISPR features (Pinilla-Redondo et al., 2019;...

...signal is in hot and the anatomical B-mode images are in greyscale. (scalebar = 2mm)   We hope you...the tumors was overlaid with the anatomical B-mode images in greyscale.   ARG expression in mammalian...production of air-filled protein nanostructures called gas vesicles, which scatter sound waves and thereby...contrast in E. coli. We deleted an unannotated hypothetical protein, improving expression, and named this...of the mARG plamsids into HEK293T cells using a chemical transfection reagent (e.g. PEI-MAX). Simply cotransfect...images of GV expressing MDA-MB-231 cells (top, scalebar = 10µm) and TEM of GVs purified from MDA-MB-231...cell lysate after 4-days of expression (bottom, scalebar = 0.5µm). (d) Ultrasound imaging of MDA-MB-231...

...said to be representative or the statistical variation is calculated and placed in the legend. Widely ...publication called Scientific Data, which aims to act as a peer-reviewed repository for “scientifically valuable... errors in assumptions, use of inappropriate statistical methods, omission of some data, etc.  The firestorm...months or years before others. In some cases, typically consortia involving genomic screens, the data ...  This means that supporting datasets must be logically annotated and arranged, as in an actual publication...published, this data is usually condensed into a statistically evaluated set of values.  However, to evaluate...likely to improve research. What is published is typically a tiny fraction of what was actually carried out...

...backgrounds with technical expertise that ranges from astronomy to bioethics to chemical engineering and...explore their Local Engagement Playbook for step-by-step guides on how to engage in your local community....societal pressures including economic, cultural, and political. Science policy looks differently depending on... p” policy.  “Big P” policy is what you might typically think of. It occurs in a government setting: drafting...in Congress, or can also happen at the state and local level. The big secret about science policy that ...STPF cohort members came to DC to do either a sabbatical year or transition away from academia as tenured...supplement to your career through engagement with your local or state governments. Most communities have opportunities...

...principles to biological systems. Today, what started in Cambridge with a handful of local high school ...BioBuilderClub teams’ biological engineering designs and ideas ranged from a biological breathalyzer to pesticide...School Apprenticeship, preparing local (Massachusetts), economically disadvantaged students for paid summer...into today’s classrooms,” BioBuilder answers the call by offering a variety of educational programs all... Dr. Natalie Kuldell from MIT’s Department of Biological Engineering. Built primarily from her 20 years...experiential learning. “Students may forget the technical details they hear in their classes,” she said,...learning outcomes by emphasizing the importance of critical thinking, persistence, teamwork, and communication...

... the so-called “reproducibility crisis.” Results in preclinical, biomedical and psychological sciences...researchers.There is both theoretical and empirical evidence (in psychology, cancer biology, pre-clinical life science...sciences were called into question after credible attempts to replicate major findings could not be replicated...practices (QRPs) can quickly lead us to find statistically significant results that are not robust. Sampling...research plans become public eventually). It is typically done right before data collection starts, and ...each hypothesis will be tested with a specific statistical test.  Preregistration makes clear the distinction...differences between groups. Hypothesis generation is critical for advancement, but we are too often encouraged...

...their visions for a successful and sustainable biomedical research sector at our first Future of Research...discussed, many related to the structure of the biomedical workforce. Quotes taken directly from participants...around the USA undertaking BEST programs, including local schools in Boston: the Boston University School ...of Medicine and the University of Massachusetts Medical School. Graduate education programs are also beginning...internship to be taken in an industry, regulatory, or clinical environment. It remains contested, however, whether...shape and inform the overall structure of the biomedical research sector, and what a tenable workforce...Shapes Science’ and contributing to the NIH’s Biomedical Research Workforce Working Group. Along with ...

...mice at a time. Because mice are more genetically and physiologically similar to humans than many other model...study calcium signaling in the entire adult brain, using a plasmid that expresses a genetically encoded... of mice as model organisms is that they are genetically tractable. Mice can be easily manipulated with...also relatively straightforward to manipulate genetically. Scientists can make a new line of flies in only...reasons above but because they actually share many biological properties and processes with our own cells. ...important experimental model in the study of neurological diseases, such as Parkinson’s disease (PD). ...embryonic development to adulthood. This ability to genetically manipulate and visualize cells and tissues allows...

...have been typically optimised for immortal/genetically-corrupt cell lines, lack scalability, and have ...alone in the cellular milieu. It is, therefore, critical to understand how proteins interact with one another... another (or with DNA) to perform diverse biochemical tasks in the cell. One of the most popular approaches... to tag the protein of interest with a small biochemical “epitope tag” such that these tagged proteins...epitope tagging in mammalian stem cells. For easy scalability to the 96-well format, we developed a “TAG-IN... purchased from various other suppliers with a typical turnaround time of 5-7 days. The crRNA and tracrRNA...optimized to achieve high knock-in efficiency. To calculate the tagging efficiency in the transfected cells...

...which aims to increase your understanding and technical proficiency with common applications - now let... differences of twofold or greater with good statistical significance (Taylor, 2017). With the right conditions...appropriate choice (Panina, 2018). Look for studies specifically aimed at determining the best options for a ...of triplicates to understand variation due to technical error.  Plate readouts can be affected by plate...sets and most machines have a protocol that automatically performs the normalization step.  Machines Most...when using fluorescent probe-based chemistry - theoretically, up to five or six reactions in a single well...and sets.   🔥Hot Tip🔥 The ABI machines automatically record fluorescent data on all channels - helpful...

...ratio calculators Lab math calculators These calculators can help you with common calculations performed...lab. Molarity calculator Promega's Biomath Calculators Primer resuspension calculator (sign-in required... Cloning ligation reaction calculators These links can help you calculate how much insert and backbone...required.)  DailyCalcs Science Calculator app for iPhone Reagent and Data Repositories These links can help...assembly, or In-Fusion cloning.  NEB Ligation Calculator: This tool works for any cloning method. You ...

...Attempts to remove excess bone surgically cause the body to “heal” the surgical site with even more bone that...therapeutic biologic design (ADCs) along with pre-clinical analytical development. His areas of expertise include...protein engineer currently working at Sanofi Pharmaceuticals in Cambridge, MA. When I joined Sanofi/Genzyme...to get in shape. After three kids, I had put on typical middle age weight and “exercise” consisted of taking... learned there was a Genzyme-based running team called Running for Rare Disease that runs on behalf of... But, there is HOPE. FOP is a genetic disease typically caused by a single R206H point mutation on the...growth in mouse models. As this biologic moves into clinical trials, there is new HOPE for AJ. But, as we in...

...ways to monitor cell, organelle, and protein localization. For instance, you might dye your cells and ...timers, add an entire new dimension to monitoring localization; using FP timers, researchers can look at a ...single image of a cell and understand how protein localization changes over time.Hallmarks of fluorescent protein...time but independent of protein concentration. Calibration curves correlating this ratio to the total expression...in whole cells or as a pool of fusion proteins localized to a specific region of the cell. For example,...fluorescence falls within the linear window for the theoretical timer shown in Figure 1B, you can use the equation...temperature, and pH, but timer activity can be calibrated to these different conditions, enabling the use...

...fluorescence microscopy vs. confocal microscopy In confocal microscopes, optical sectioning of a specimen...sheet, typically 2-6  microns, illuminates only one thin plane of the sample surrounding the focal plane...Lee and Pantelis Tsoulfas of the Department of Neurological Surgery at the University of Miami. The beginning...expressing eGFP under the human ubiquitin C promoter. Anatomical abbreviations: Gr= Gracile tract, Cu= Cuneate...originally named orthogonal-plane fluorescence optical sectioning (OPFOS) (2). Arne Voie, David Burns ...illumination (i.e. light sheet).  This is in contrast to confocal laser microscopy where the laser and the reflected..., the photomultipliers used to detect light in confocal microscopes are less efficient than the modern...

...visualize the localization of the FP (fused to the protein of interest), thus tracking the localization of that.... The most commonly used are GECIs - genetically encoded calcium indicators (e.g. GCaMP). Others include...capacity. Many of these FPs are fluorescent when ectopically expressed in most organisms. Furthermore, fusing...fluorescence. Therefore, FPs are used to study many biological questions. The two most common uses are: 1) to...following their activation. The four main FTs are called Slow-FT, Medium-FT, Fast-FT, and mK-GO. Large Stokes...For LSS proteins, the Stokes shift is ≥ 100nm. Specifically, these proteins are excited by UV light or blue...of the split FP. This technique is also is also called Bimolecular Fluorescence Complementation (BiFC)...

...the sandwich and typically takes less than an hour. However large proteins typically do not transfer as... boiling in the presence of a reducing agent, typically containing thiols, to cleave disulfide bonds. ...gel on top of the resolving gel. A stacking gel typically has a different ionic strength and lower pH and...nitrocellulose or polyvinylidene difluoride (PVDF) in a step called protein transfer. While both nitrocellulose and...are “sticky” and have the potential to bind nonspecifically to the antibodies used for staining. To address... the controls listed on the vendor’s website. Typically, the vendor will demonstrate that their antibody...binds to endogenously expressed protein at physiologically relevant levels. Some vendors will also test...