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We narrowed to 839 results for: IGH@

Showing: 81 - 120 of 839 results

  1. Hot Plasmids - March 2020 - base editors, GEVI, MoClo, and optogenetics

    Type
    Blog Post
    Published
    March 3, 2020, 2:15 p.m.
    ...response and brightness. They also do not spectrally separate from green sensors and blue-light-activated...Physiol 2018. PubMed PMID: 30819783. Light up CreLite: a red-light inducible optogenetic tool for precision...Every few months we highlight a subset of the new plasmids in the repository through our hot plasmids...fluorescent protein (mScarlet) with either a seven or eight amino acid linker between them. The Ace-mScarlet...potential, Ace absorbance increases and results in higher quenching of mScarlet emission. Figure 2... with existing green-fluorescent GEVIs and blue-light-sensitive channelrhodopsins. Find the Ace-mScarlet...optogenetics. Cre activity is controlled by red light by splitting Cre and fusing each half with PhyB ...

  2. Hot Plasmids May 2018 - Optogenetics, Decaffeination, Biosensors, and Fluorescent Protein Tools

    Type
    Blog Post
    Published
    May 15, 2018, 12:43 p.m.
    ...them to interact by stimulation with blue light.  Similarly, light can also be used to control homo-oligomerization...seconds after a pulse of blue light, and is reversible upon termination of light stimulation. CRY2clust enables...faster assembly and disassembly dynamics, higher sensitivity to light, and better distribution of clusters ...Every few months we highlight a subset of the new plasmids in the repository through our hot plasmids...oligomerization of target proteins in response to blue light. This tool, CRY2clust, was created by adding a 9...from Arabidopsis thaliana. When exposed to blue light, CRY2 undergoes a conformational change that permits...drawback of this method is the stray excitation of neighboring neurons in opsin dense regions. To overcome this...

  3. Plasmids 101: Optimizing Plasmid Yields

    Type
    Blog Post
    Published
    May 26, 2016, 2:30 p.m.
    ...to increase plasmid yield. Keep in mind that you might need to use multiple techniques for a particularly...additional steps are needed to optimize yield. You might want to include an extra wash step or use a different...recommends DH5𝝰! This strain consistently gives good, high quality preps due to its endA1 recA1 relA1 genotype...repeats. We use NEB Stable to transform and prep high quality DNA that contains these elements. Be aware...wash step to remove endonucleases (see figure at right for an example). Look out for carbohydrates Certain...purification use the cell’s resources to produce high levels of protein, not plasmid DNA. If you need ... increasing the culture volume, be aware that a higher volume of lysate passed over the binding matrix...

  4. Sensing Neuronal Dopamine

    Type
    Blog Post
    Published
    July 30, 2019, 12:59 p.m.
    ...of the sensor, dLight1.1 and dLight1.2, exhibit high (submicromolar) DA affinity and high-magnitude responses...(Figure 2). The dLight1 dopamine sensor In 2018, Lin Tian’s lab has developed dLight1, a molecular tool... of DA release.  Using dLight1 sensors in experiments The Tian lab used dLight1 both in brain slices and...Feelings of delight associated with the brain’s reward circuits are generally linked to dopamine (DA)...Similar to calcium and glutamate sensors, the new dLight1 sensor is a single fluorescent protein-based sensor...learning and reward consumption in live mice Find dLight sensor here! The GRABDA dopamine sensor Yulong ...binding (Sun et al., 2018). These sensors also exhibit high magnitudes of fluorescent responses. Studies on ...

  5. 27 Hot Plasmids from 2016

    Type
    Blog Post
    Published
    Dec. 22, 2016, 3:03 p.m.
    ...Every quarter we highlight a subset of the new plasmids in the repository through our hot plasmids articles...articles. These brief articles highlight the main features and applications of a partiular plasmid or ...parallel analysis that successfully identified new, highly specific agonists for orphan GPCRs demonstrating...are found in one third of all cancers, including a high percentage (up to 95%) of pancreatic cancers. Ras...an additional 6 gRNAs per gene generated using slightly relaxed gRNA design parameters. To demonstrate...method in bacteria that enables rapid, automated, and high-throughput genome editing is multiplex automated...otherwise unmodified bacterial strains (5). MutL is highly conserved in distant relatives of E. coli so it...

  6. Hot Plasmids - August 2020

    Type
    Blog Post
    Published
    Aug. 18, 2020, 1:15 p.m.
    ...sometimes these fluorescent proteins might not localize to the right places, which can provide misleading...Every few months we highlight a subset of the new plasmids and viral preps in the repository through ...efficiency of tobacco etch virus protease (TEV), a highly sequence-specific protease used to cleave affinity...work for evolution of other target proteins and might be useful for your studies.  Find the plasmids to...including fusions with Histone 2A or 2B can result in bright targeted expression while minimizing non-specific...microtubules of mammalian cells, the author found that too high expression of these markers can lead to non-specific...biotin and biotin-4-fluorescein to be dissolved at a higher concentration, and results in uniform NB biotinylation...

  7. Transferable Skills Guide: Problem Solving

    Type
    Blog Post
    Published
    Nov. 30, 2021, 2:15 p.m.
    ...Addgene, we've set out to fill this gap by both highlighting that researchers do learn MANY transferable ...the lab and by offering advice on areas where you might need some help. Today in our transferable skills...strategy to fix the issue. I was problem solving!  You might think that being able to figure out what went wrong...skills” and look at how scientists can develop and highlight these valuable skills. Hopefully, having a better...everything or revealed another issue in the process, you might need to go back to the beginning, reassess the situation...is just a question you have about the world. You might do some background research to better understand...issue. Depending on the particular problem, you might even be expected to write up a report to communicate...

  8. CRISPR 101: Cytosine and Adenine Base Editors

    Type
    Blog Post
    Published
    Feb. 13, 2025, 2:15 p.m.
    ...publication highlights In the table below, you'll find a list of publications we've highlighted in this article...Publication Plasmids   Highlights Komor et al., 2016 BE1, BE2, BE3   BE3 displays highest editing efficiency...the lab created HF-BE3, a base editor containing high fidelity Cas9 variant HF-Cas9 (Rees et al., 2017...fold less off-target editing than BE3, with only a slight reduction in on-target editing efficiency. To further... positions 4-7. ABEs 6.3, 7.8, and 7.9 display slightly wider editing windows of position 4-9, although...positions A3-A4 and A8-A10 at NGG PAM and 4.2-fold higher editing efficiency at non-NGG PAM variants compared... window of only a few nucleotides, so they are tightly constrained by PAM availability in the genome. ...

  9. Technique: Probe Phage Genomes for Host Binding Proteins

    Type
    Blog Post
    Published
    Sept. 29, 2016, 2:30 p.m.
    ...using a highly competent strain that you can use for long-term library storage. After overnight growth,...popular classes of phage proteins to study. These are highly useful for the biotech industry, which is in the...the plate so you can regrow the lifted colonies overnight), flip the membrane over and place it colony-side...tested, as with any protein purification, but overnight at 30°C is a good start. Prepare a lysis solution...repeated 5 times) works well. Leaving the membrane overnight in the freezer during one of these steps is also...with your cells of interest and growing them up overnight. Assuming they expressed well, the proteins encoded... it can really help to incubate the membranes overnight in 500 mM to 1 M NaCl in order to enhance folding...

  10. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

    Type
    Blog Post
    Published
    Feb. 4, 2020, 2:15 p.m.
    ...color: they strongly absorb visible light to give colors in ambient light. In contrast, the widely used green...protein (GFP) gives a bright green fluorescence when exposed to blue to ultraviolet light, whereas chromoproteins...chromoproteins absorb visible light and give off color in ambient light, it gives scientists the ability...cases, cells containing high-copy plasmid-borne chromoproteins lost color overnight in liquid culture, though...colorful corals, to name a few. What’s behind these sights are an abundance of biological molecules like chromoproteins...Cassiopea xamachana (Blanquet and Phelan, 1987). Eight years later, scientists isolated pink, purple, and... irradiation by reflecting visible and infrared light to dissipate excess energy at wavelengths of low...

  11. Better Dyeing Through Chemistry & Small Molecule Fluorophores

    Type
    Blog Post
    Published
    Sept. 8, 2016, 2:30 p.m.
    ...substantially brighter and more photostable than fluorescent proteins, providing a straightforward way to improve...refinements to these systems—smaller tags, brighter conjugates, higher fluorogenicity, photoactivation etc.—...fluorophores conjugated to molecular species with high affinity for endogenous molecular targets such as...selectivity of the fluorophore attachment, the brightness of the resulting conjugate, and the complexity...exhibits low fluorescence when free in solution, but high fluorescence upon binding to its cognate protein...due to its established chemistry, small size, brightness, and cell permeability. In past decades, further... the Alexa Fluor and ATTO dyes, with improved brightness, photostability, and spectral range. Nevertheless...

  12. Hot Plasmids September 2018 - Optogenetics, RNA Localization, Fluorescent Protein, and Base-editing Tools

    Type
    Blog Post
    Published
    Sept. 18, 2018, 1:33 p.m.
    ... Zdk1 binds LOV2 with high affinity in the dark but dissociate under blue light. This tool has now been...Every few months we highlight a subset of the new plasmids in the repository through our hot plasmids...up here.   Listen this episode of Hot Plasmids! Light controls the activity of a microtubule-associated... lab at UCSF to modulate cellular dynamics with light by modifying EB1. EB1 normally sits on microtubules...microtubule growth can be switched off with blue light which causes LOV2 and Zdk1 to dissociate due to ...photodissociable EB1 could be stopped in areas with blue light - a finding which opens novel ways in manipulating...Scientists are always on the lookout for better, brighter fluorescent proteins (FP) with improved tissue...

  13. Hot Plasmids - June 2020 - Barcoded CRISPR Library, Sparse Cell Labeling, Calcineurin Reporter, and DNA Staining Dye Alternative

    Type
    Blog Post
    Published
    June 2, 2020, 1:15 p.m.
    .... The iBAR method resulted in high quality data even when used at a high multiplicity of infection, because...also function in high salt conditions - up to 100 mM NaCl. These tTALE-FPs also have higher DNA binding affinity...Every few months we highlight a subset of the new plasmids in the repository through our hot plasmids...next generation sequencing.  In order to avoid a high false-positive rate discovery rate, conventional...human gene, it enables genome-wide screening with a high MOI, which is beneficial for a low number of cells... levels within the cell. When levels of Ca2+ are high in the cell, calcineurin dephosphorylates the protein... the bacterial βgal assay, and are not suited to high throughput screening, calcineurin dynamics measurements...

  14. Extracurricular Activities for a Strong Science Career Path

    Type
    Blog Post
    Published
    Jan. 21, 2014, 3:08 p.m.
    ...development. However, careers outside of the lab highlight “transferable” skills such as leadership, management... impact, or a journalist does not understand the high-level science discussed in an interview, your project...be taboo in the lab, multiple research studies highlight the importance of taking a break and incorporating...network Attend informational interviews to gain insight into the daily routine of the potential job Join...and research 2 – 3 potential professions will highlight which skills you should focus on improving and... skill sets are strengthened. This short list highlights the ubiquity of skills across professions within... Outreach activities (e.g. Volunteer at a local high school science class: Science Club for Girls) ...

  15. Plasmids 101: Codon usage bias

    Type
    Blog Post
    Published
    Sept. 27, 2018, 1:09 p.m.
    ...amino acids (see example to the right). With such simple rules, you might think it’s easy to come up with... entire translation process. For example, during high growth rate conditions, E. coli preferentially upregulates...production of tRNAs that recognize codons found in highly expressed genes (Emilsson and Kurland, 1990). Controlling...produced at a lower rate than proteins encoded by highly abundant, charged tRNAs. For example Tuller et ... protein is encoded by a mixture of codons with highly and poorly charged tRNAs, different regions of ...usage, an organism can change which proteins are highly expressed and which are poorly expressed by producing... during amino acid starvation thus resulting in higher production of amino acid biosynthetic enzymes (...

  16. Plasmids 101: Plasmid Incompatibility

    Type
    Blog Post
    Published
    March 31, 2020, 1:15 p.m.
    ...cell is artificially high for both plasmids, so each plasmid “thinks” it has a higher copy number than it... 101 post, we’ll cover why this happens, how it might affect your work, and how understanding it can be...inhibits replication when copy number in the cell is high, but allows replication when copy number in the ...replication machinery when plasmid copy number is high (del Solar et al., 1998).  In cases of compatible...plasmids, though the partitioning problem does occur in high copy plasmids (Diaz et al., 2015). Plasmids contain... plasmid pairs into the new daughter cells.  For high copy plasmids, incompatibility due to partitioning...incompatibility due to having the same replication machinery. High copy plasmids with the same centromere-like binding...

  17. Tips and Tricks for Using Golden Gate Modular Cloning (MoClo)

    Type
    Blog Post
    Published
    July 8, 2025, 1:15 p.m.
    ...Save time, go high throughput You are ready to perform your assemblies, but you have a high number to get...E. coli strains with higher transformation efficiency. If you’re cloning in high-throughput, we recommend...ligation efficiency, we recommend you calculate the right volume of each plasmid according to their concentration...time to think big and perform your assemblies in a high-throughput format. If your lab owns a liquid handling...restriction–ligation reactions in a 96-well (or higher!) format. Liquid handling robots are not only faster...experimental costs. If you are planning to perform a high number of assemblies, you will also likely need ...your bacterial stocks beforehand. You can also use high-throughput methods to streamline your plasmid extractions...

  18. Fluorescent Protein Travel Awards - FLiPs and Fluorescent Protein Biosensors

    Type
    Blog Post
    Published
    July 27, 2017, 1:11 p.m.
    ...aggregation tendency, increase its brightness, and decrease the amount of light required for dimerization. She...Fluorescent Proteins Use Light to Control CRISPR Genome Editing Choose the Brightest Fluorescent Protein for...future award opportunities. Xin Zhou: Fluorescent Light-inducible Proteins (FLiPs) Fluorescent proteins ...create enzymes that can be turned on and off with light.  Xin hypothesized that she could make enzymes controllable...known to oligomerize when exposed to ultraviolet light. To do so, she generated fusion proteins in which...enzyme’s active site. In these fusions, ultraviolet light exposure caused oligomerization of the Dronpa domains...shutting off the enzyme. Later exposure to cyan light stopped Dronpa oligomerization and turned the enzymes...

  19. Choosing Your Fluorescent Proteins for Multi-Color Imaging

    Type
    Blog Post
    Published
    Oct. 9, 2014, 3 p.m.
    ...some light on multicolor imaging with fluorescent proteins. With the right microscope and the right choice...when choosing an individual fluorescent protein (brightness, photostability, and so on; see the previous ...colors and the mCherry spectrum is shown in the lighter colors; excitation spectra are blue and emission...this filter set, we would expect TagRFP to give a brighter signal than mCherry. In general, filter sets designed...supplement your cells with biliverdin for maximal brightness. In mammalian cells, one of the improved folding...fluorescent protein that is supposed to be extremely bright. In S. cerevisiae, we've tested a number of green...proteins with this filter set and have reported brightness measurements. Here, EGFP outperforms the improved...

  20. Viral Vectors 101: Types of viruses

    Type
    Blog Post
    Published
    June 6, 2023, 1:15 p.m.
    ...are commonly used in the lab as tools and we all might benefit from getting to know them a little better... injury, retroviruses generally have moderate to high immunogenicity, meaning they induce an immune response...transduce (infect) a wide range of cell types with a high efficiency. However, gamma-retrovirus infection ...tools have been derived from. Lentiviruses have slightly larger packaging capacities than gamma-retrovirus...immunogenicity, meaning it does not induce the same high immune response as retroviruses. This virus can ...been identified so far! Adenovirus generally has high immunogenicity, but some strains have been engineered... discussed in this article, adenovirus has the highest maximum titer, a plus for production and infection...

  21. Transferable Skills Guide: Time Management

    Type
    Blog Post
    Published
    Dec. 5, 2017, 1:52 p.m.
    ... university. My to-do list might look something like the image on the right (and for those of you who ...Addgene we've set out to fill this gap by both highlighting that researchers do learn MANY transferable ...the lab and by offering advice on areas where you might need some help. Today in our transferable skills...tasks at the expense of time spent on the long and highly useful tasks. The quick accomplishments will likely...you the energy to finish any additional work you might need to do. In most cases, I would recommend the...constantly switching between different projects, you might not be able to make any reasonable progress on any...to-do lists at the end of the day so they can jump straight into tasks the next day. If you work in a job ...

  22. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

    Type
    Blog Post
    Published
    April 24, 2015, 2:39 p.m.
    ...ideal characteristics (low aggregation, high photostability, and high stability after fixation.) From the ...still being developed today. In this post, I will highlight the evolution of the  Brainbow multicolor labeling...zebrafish and Drosophila. Why Brainbow? Brainbow highlights the trend in biology towards single-cell resolution...development of a new technique. In a structure as tightly packed and interwoven as the brain, robust single-cell..., one would expect ten colors (see table to the right); in various reports, Sanes and Lichtman have observed...observed from 90-160 distinct colors due to higher copy number. The name Brainbow is truly fitting for...fluorescently distinct, their protein sequences are highly homologous, preventing design of antibodies specific...

  23. Antibodies 101: Fab Fragments

    Type
    Blog Post
    Published
    Sept. 5, 2024, 1:15 p.m.
    ...bivalent, so may have higher avidity than monovalent Fab’ or Fab fragments. Higher avidity means that F...’ll go over what a Fab fragment is and when you might use it in the lab. What is a Fab Fragment? Consider...chains (two identical heavy chains and two identical light chains) each with a constant region and a variable...heavy chains and between one heavy chain and one light chain. When assembled, the four chains make a Y-...are the “arms” of the Y and comprise the entire light chain and a variable and constant region of the ...important features labeled. V = Variable domain. L = Light chain. C = Constant domain. H = Heavy chain. Created...function but not the Fc region (more on when that might be later). Find scFv plasmids at Addgene Depending...

  24. Developing Lab Management Software for Biology

    Type
    Blog Post
    Published
    Aug. 25, 2016, 2:30 p.m.
    ...the contents. Tightly coupling barcodes and containers might not be such a bad idea, tightly coupling them...we’ll highlight some of the lessons we’ve learned over the years with the hope that our insights can help... help steer you in the right direction when writing your own software. None of this is gospel, but we ...help tell this story. In this first example to the right (# this will make you sad),  someone has put in ...numbers. Loosely couple containers and contents Right from the get-go be as clear as you can be (there...them to their contents might leave you sad and confused when a tricky procedure failed and one of your...In the first example we’ve used inheritance to tightly couple the contents of a tube with the tube itself...

  25. Finding Your Perfect Job After University

    Type
    Blog Post
    Published
    Jan. 12, 2016, 3:30 p.m.
    ...only scientists with an MSc or higher apply. It was important to weigh the cost to benefit of studying... I didn’t need to explain why I changed jobs. I highly recommend taking on a similar position in your ...your field of interest before you dive straight into a full-time position. For my part, I really enjoyed ...for a disease that traditional medicine couldn’t fight effectively. I had to learn how to cope with a different... companies which are rapidly expanding, as this might not be sustainable. After working at this company... of different types of pancreatic cancer and highlighting which treatment would produce the best outcome... does not work for all patients, leading to very high mortality. I really loved the fact that my work ...

  26. A Look at Addgene's QC Process

    Type
    Blog Post
    Published
    May 6, 2025, 1:15 p.m.
    ...error — both human and biological. Even the most high-fidelity polymerases can make mistakes and lead ...these errors. These processes allow us to have a high standard of sequence quality across all plasmids... same QC process to ensure all plasmids pass our high-quality standards and contain the expected plasmid...accurate The gene/insert or any major feature highlighted by the deposited scientist  The backbone elements...deposited to Addgene. This review process also highlights other QC topics that may require additional attention...Illumina MiSeq platform, such as GC-rich regions and highly repetitive sequences and/or features. Most of these...assembly in Geneious can still provide additional insights to resolve the assembly and determine if there...

  27. Fluorescent Proteins 101: History of Fluorescent Proteins

    Type
    Blog Post
    Published
    Aug. 7, 2017, 1:58 p.m.
    ...easily detect and measure the light they emit. Proteins that give off light include chemiluminescent proteins...victoria there is a ring that glows with a faint green light. To investigate this green glimmer, Shimomura collected...which he called aequorin, gives off a faint blue light when it binds calcium ions, and the second, which...which we now call GFP, absorbs that blue light and glows green. In the late 1980s, another researcher, Douglas...protein localization. And as it turns out, he was right! He began to study the A. victoria gene responsible... fell into place, as they, taken alongside the insights provided by the studies in Tsien lab, covered ...their work on GFP. This great honor serves to highlight how useful fluorescent proteins like GFP really...

  28. Let There Be LITE Plasmids

    Type
    Blog Post
    Published
    Nov. 21, 2013, 3:56 p.m.
    ... and Mark Brigham have taken the technology to another level with the addition of two light-sensitive ...occurs within minutes of light exposure. Third, the response can be graded with light dose and is fully reversible...reversible after light retraction. Finally, because light can be applied non-invasively, its use is not...borrowed from Arabidopsis thaliana. The results are light-inducible transcriptional effectors (LITEs) designed...them on or off, literally at the flip of a (blue) light switch. Zhang’s team devised the plasmids now available... transcription in as little as 30 minutes after light stimulation. Under those optogenetic controls, gene...several advantages of the new system: "First, the light-responsive molecules of cryptochrome 2 are the chromophores...

  29. PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs

    Type
    Blog Post
    Published
    June 25, 2024, 1:15 p.m.
    ...understand how chromatin might influence editing results at your target. Figure 3: High-throughput screening...comes in! Evaluate prime editing efficiencies in high-throughput libraries My labmates and I wanted to...of pegRNA designs based on patterns observed from high-throughput experiments, where we can evaluate tens...of designs in parallel. To do this, we performed high-throughput ‘self-targeting’ screens with oligo libraries... edit types and cell contexts. Figure 1: High-throughput screening of pegRNA efficiency and building...screens such as PRIDICT/PRIDICT2.0 provide valuable insights into pegRNA editing efficiencies for specific ...research by the Shendure lab (Li et al., 2024) have highlighted how chromatin substantially affects prime editing...

  30. Deep Dive: Statistical Tests (Comparisons)

    Type
    Blog Post
    Published
    March 7, 2023, 2:15 p.m.
    ...needed, without a high number of samples (~5). Tukey’s test – Most conservative test. Higher false negatives...select the right test for the job (and, uh, gentle reminder: keep your friendly neighborhood biostatistician... can tell you, and how to identify which one is right for you. Statistics – The basics Statistical analysis...ANOVA. Isn’t that what a two-way ANOVA test is you might ask? Not exactly! A MANOVA test assesses the effect...could test the impact of temperature, humidity, sunlight exposure time, and watering volume on total crop...total), your likelihood of a false positive is much higher than if you only look at 2 or 3 pairwise comparisons...range test – Most powerful. Low false negatives, high false positives. Control vs. all samples Oftentimes...

  31. Antibodies 101: Antibody Engineering and Directed Evolution

    Type
    Blog Post
    Published
    Aug. 5, 2025, 1:15 p.m.
    ...even different chimeras might function better. In that case, gene shuffling might be a good option. In gene...random library of mutants using error-prone PCR might be a good solution. The process of error-prone PCR...you wouldn’t want errors in a PCR and would use a high-fidelity polymerase to ensure accuracy. In this ...more control, structure guided directed evolution might be more your style. Instead of leaving sequence ...regions you want to evolve. With this added layer of insight, you can shrink your mutant library and increase... of directed evolution, all while maintaining a higher degree of control and efficiency! Screening mutant...improved properties across large mutant libraries, high-throughput screening techniques such as ELISA and...

  32. Don’t FRET: Bimolecular Fluorescence Complementation Makes Visualizing Protein-Protein Interactions Easy

    Type
    Blog Post
    Published
    Aug. 27, 2020, 1:15 p.m.
    ... the assayed PPI. This high-throughput application is much more straightforward to carry out because it... This is straightforward in easy-to-transfect cell lines where plasmid dosage can be tightly controlled...the non-radiative transfer for energy between neighboring fluorophores, FRET can be used to detect the ...subcellular localization of the studied PIPs. The straightforward nature of BiFC also makes it easier to generate...generate a large number of constructs for high-throughput screening of PPIs. For example, Bischof et al....Hu, 2012). Having a rough idea of how your PIPs might interact will allow you to determine where to fuse... Proteins 101: GFP Fusion Proteins - Making the Right Connection. Cellular expression Whether BiFC constructs...

  33. Getting to Know Expansion Microscopy

    Type
    Blog Post
    Published
    June 20, 2023, 1:15 p.m.
    ...common reagents, straightforward protocols, and standard microscopes sounds great, right? But there is always...         Why Expansion Microscopy?  Traditional light microscopes are limited in the level of resolution...everything needed for synaptic transmission, are right around 250 nm, making it difficult to study the ...the inner workings of the synapse by conventional light or fluorescence microscopy (Dani, et al., 2010)....What is Expansion Microscopy?  ExM provides a straightforward answer to the space problem. Instead of imaging..., biological samples are enmeshed in a gel of a highly absorbent material that is able to expand evenly...points with a conventional, diffraction-limited light microscope.  Along with using more accessible imaging...

  34. Plasmids 101: Choosing an Antibiotic Resistance Gene

    Type
    Blog Post
    Published
    Oct. 10, 2023, 1 p.m.
    ... colony formation on agar plates, less stable at high temperatures and in acidic environments.   Carbenicillin...better’ antibiotic than amp: it is more stable in high pH environments and is less susceptible to degradation..."fontawesome-5.14.0" name="Check" style="SOLID" height="16" purpose="decorative" title="Check icon" %}...

  35. SunTag and Fluorescent Imaging

    Type
    Blog Post
    Published
    March 28, 2017, 2:30 p.m.
    ...protein at higher levels, you risk observing artifacts only present when the protein is at a very high concentration...membrane-targeted CAAX-SunTag was 18-fold brighter than sfGFP! The high SunTag signal allowed them to cut their...transcription factor binding sites in a promoter lead to higher transcriptional activation. Multiple nuclear localization...explosion SUperNova," SunTag can help you turn up the brightness in your fluorescent imaging experiments. Fluorescent...their laser power by over 80% and still obtain a higher signal-to-noise ratio with a lower photobleaching...mitoNEET-SunTag-GFP. As expected, they obtained bright images of mitochondria without organelle toxicity...to traditional fluorescent proteins Improved brightness and signal-to-noise ratio Less chance of phototoxicity...

  36. Deciphering the Mysteries of Behavior with Viral Vectors

    Type
    Blog Post
    Published
    Sept. 17, 2019, 12:40 p.m.
    ...elevated plus maze, the light-dark box is a simple device comprised of a brightly lit (risky) and a dark... a camera or tracking software. Locomotion is a highly complex process on the level of individual neurons...findings by controlling neurons in the mPFC with light. Activation of these neurons via the channel Channelrhodopsin...inhibitory mechanism of locomotor sensitization and might open new ways for a novel therapeutic approach against...to stay in dark areas (protected) as opposed to brightly illuminated ones (risky). They also prefer to ...diseases, animals have reduced anxiety levels and a higher tendency for risk taking. These animals also show... arm (risky), or a closed arm (safe). Mice with higher IL-1 beta production spent more time in the open...

  37. The Strength of Story Telling

    Type
    Blog Post
    Published
    Dec. 19, 2017, 2 p.m.
    ...make sure people pay attention. Highlight your lowest points and highest points and make clear what was...I’m working with a talented journalist to shine light on living as a scientist in Beijing, talking to ...judgement. Eventually, we published the paper in a high-impact journal and it worked out for all of us.”...cases, the protagonist will be you. Make sure to highlight what lessons you’ve learned and what knowledge... into your research at an outreach event as you might in a scientific paper. Build lots of versions of...difficult and what was easy when appropriate. It might not be scientific to bring emotional discovery into...people pay attention. Practice Nobody gets good overnight. The only way to improve is to keep working on...

  38. Hassle-free 96-well Format Epitope Tagging Using Cas9 Ribonucleoprotein

    Type
    Blog Post
    Published
    June 28, 2018, 3:01 p.m.
    ... is the cr:tracrRNA approach highly efficient, it is also straightforward as the RNA molecules can be ...events. In the light of these results, we re-analyzed our data and found that 75% of our highest knock-in efficiency...of interest are pulled-down and later coupled to high-throughput sequencing to identify genome-wide binding...Telmer 1998). Indeed, this method can be used at high throughput because numerous cell lines can be generated...challenges, we set out to develop a hassle-free, highly efficient Cas9 protein-based method for epitope...which, in our hands, is both cost-effective and highly active. After csRNP complex formation, a single-stranded...transfection/delivery step needs to be optimized to achieve high knock-in efficiency. To calculate the tagging efficiency...

  39. Special Delivery: Fluorophore Targeting for FRET Studies

    Type
    Blog Post
    Published
    July 19, 2016, 2:30 p.m.
    ...dye-nitrilotriacetic acid (NTA) conjugates (Kapanidis, Ebright, & Ebright, 2001). These Cy/NTA conjugates work the ...6. Kapanidis, Achillefs N., Yon W. Ebright, and Richard H. Ebright. "Site-specific incorporation of fluorescent... James D. Fessenden, an Assistant Professor at Brigham and Women’s Hospital. Biochemists often struggle...mailbox: How to get your fluorophore delivered to the right place? FRET experiments require targeting the donor...the proper mailbox (i.e., a binding site) in the right city (protein). In a cell, this task is even harder...acceptor molecules are targeted nearby (such as a neighboring house on the same street as the donor), then ...specificity Donor fluorophores need to go to the right place. Mis-targeted donors never “see” FRET acceptors...

  40. Plasmids 101: Expressing shRNAs for RNA Interference

    Type
    Blog Post
    Published
    June 17, 2025, 1:15 p.m.
    ... handy “oldie” as you might say. RNA interference (RNAi) is, as the name might suggest, an RNA-mediated...Addgene plasmid #11653). Full shRNA sequence is highlighted in green, with red boxes indicating mature siRNA...Addgene plasmid #21114). The miRNA sequence is highlighted in green, with red boxes indicating mature miRNA...miRNA. Hairpin loop is in between the red boxes. Highlighted blue regions show flanking genomic sequence. ...-radius":"10px","padding-left":"180px","padding-right":"180px"}{% endraw %}{% end_module_attribute %}{...font-family":"helvetica","font-style":"normal","font-weight":"normal","text-decoration":"none"},"fallback":...font-family: helvetica; font-style: normal; font-weight: normal; text-decoration: none","font_set":"DEFAULT...
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