We narrowed to 886 results for: URE
-
TypeBlog PostPublishedMarch 24, 2016, 2:30 p.m....strand in Figure 1 below) was free to anneal to complementary ssDNA molecules in vitro. Figure 1: Cas9...presumptive mechanism of action are presented in Figure 2. Figure 2: Single-stranded donor DNA interaction...Understanding Cas9 biophysics enhances HDR We first measured Cas9 dissociation rates and discovered that the... of approximately 6 hours. To our surprise, we measured identical dissociation rates for catalytically...catalytically inactive dCas9 produced a “bubble” structure in uncut target DNA that was accessible to anneal...single stranded DNA. We thus wondered if such a structure could drive sequence replacement in cells. Targeting...inactive CRISPR-Cas9 using asymmetric donor DNA." Nature biotechnology (2016). PubMed PMID: 26789497. 2...
-
Management for Scientists: Delegating is Key
TypeBlog PostPublishedFeb. 20, 2015, 8:30 p.m....make sure the work gets done and done well. Involving your team effectively is now your new measure of ...Setting someone up for failure by delegating is a bad idea. How do I make sure it goes well? As with ...appropriate communication between members of the team to ensure that no component of the overall project gets left...fun, but getting buy-in when delegating can help ensure successful results. Can you provide the information...will be made that could have been prevented. Make sure everyone understands that no question is “dumb” ...and plan changes. Written documentation helps make sure everyone is on the same page. Effective delegation... the best use of your time and skills. It also ensures that other people in the team grow and develop ... -
A Practical Approach to Choosing the B(right)est Fluorescent Protein
TypeBlog PostPublishedJune 1, 2017, 2:30 p.m....brightness in a system closely mimicking the future application. In figure 2 we provide an example of the comparison...spent and, if you do it right, can be (part of) figure 1 of your next paper or thesis. This series of ...protein is based on the properties found in the literature or tabulated data that summarizes properties ... and therefore high brightness is an important feature of a fluorophore. The key question is how the brightness...detector sensitivity) and the biological system (temperature, prokaryote versus eukaryote, background fluorescence...within the dish. This is caused by the stochastic nature of plasmid uptake by cells. To correct for this...2010). An explanation of the assay is depicted in figure 1. The strict correlation between the fluorescent... -
How Addgene Creates Content
TypeBlog PostPublishedNov. 17, 2022, 2:15 p.m....process led to quick dives back into the literature to make sure absolutely everything was correct. Final... we usually make figures in BioRender or use appropriately licensed and cited figures from papers or image...content - the useful information on scientific literature, technical skills, and experiences from thousands... Nearly everything we write entails a primary literature search to collect up to date information on the...Final touches We try to use figures and images to help illustrate our points and break up walls of text...graphic designer made most of the Antibody Guide figures. This requires a fair amount of communication between...our content team and our graphic designer to make sure things are clear, accurate, and in line with Addgene... -
Antibodies 101: ChIP
TypeBlog PostPublishedMarch 10, 2022, 3:25 p.m....interactions. ChIP works by using antibodies to capture protein:DNA complexes with antibodies specific ...your protein(s) of interest. Once the complex is captured, it is treated so that the protein and DNA are...Once you have your sample, the first step is to ensure your protein:DNA complexes will remain bound to...size, typically between 200-1000 bp. This is to ensure that your downstream analysis focuses on the DNA...the sample, introduces bias due to the selective nature of the cutting. While sonication is usually the...DNA and then determine concentration. Protein Capture After crosslinking and sonication or enzymatic ...native proteins (so antibodies validated only for denatured proteins may not be a good choice) and the amount... -
Advice for Starting a Biotech Company from a Grad Student turned Entrepreneur
TypeBlog PostPublishedJuly 10, 2014, 7:17 p.m.... unable to secure enough money to sustain a startup, he went to work for Flagship Ventures – a local Cambridge...in academia and industry vary drastically, he reassures that one can learn it by reading a broad variety...of materials and constantly practicing. “The expenditure of effort in preparation and practice is directly... Cambridge biotech venture capital firm with deep ties into the local academic community. However, Mike...entrepreneur must be OK with the constant fear of failure at all stages in the startup". To gain insight ...and vision for the company; this will help avoid future complications and potential conflicts that waste...takes around 1 – 2 years. The product is the measure of success. Technology is proven in (A) Productize... -
Viral Vectors 101: AAV Variables That Matter
TypeBlog PostPublishedJune 1, 2023, 1:15 p.m....about are (1) your experimental conditions and measurements; (2) the cells you’re targeting; (3) how much...vector: serotype, promoter, and titer. Figure 1: Flow chart depicting a) steps for selecting ...injection site than AAV2 (Watakabe et al., 2015) (Figure 1). You’ll need to consider if such properties ...goals when designing and testing your AAV. Figure 2: Comparison of AAV serotypes 1, 2, 5, 6, 8, and...regions. Image modified from Aschauer et al., 2013, Figure 1. Used under Creative Commons Attribution License... 2004; Taschenberger et al., 2017). Figure 3: Comparison of two astrocyte selective promoters...neurons (NeuN). Image from Griffin et al., 2019, Figure 4. Used under Creative Commons Attribution 4.0 ... -
Recombinase-based State Machines Enable Order-dependent Logic in vivo
TypeBlog PostPublishedJuly 28, 2016, 2:30 p.m....to the system and in what order they were added. Figure 1D below demonstrates an example of a register ... in certain states of interest. For example, in Figure 2 below we present a two-input, five-state GRSM...we implement RSMs in E. coli with two plasmids (Figure 3 below): Input Plasmid: The input plasmid (high... RSM (See performance data from the indicated figures in our paper (1)) Input Plasmid Output Plasmid ...designed to have 20-30 nt length, ~60 oC annealing temperature, and ~50% GC content) so that individual parts...Ara) RSM over a time series of increasing input exposure lengths and found that two hour inductions were...each possible permuted substring of inputs (see Figure S9 of our paper (1)). We anticipate that, depending... -
Troubleshooting Your Plasmid Cloning Experiment
TypeBlog PostPublishedSept. 24, 2019, 12:55 p.m....are doing multiple digestion be sure that the buffers and temperature are compatible between the different...or use a different temperature (30°C) for the growth on plates and in liquid culture. If you don’t get ...mindful of the restriction enzymes you chose. Make sure that enzymes that you are using are not blocked ... few general parameters that you can adjust: temperature and time of incubation, and amount of DNA. As...a few colonies (5-10) and grow them in a small culture volume (2 or 3 ml) containing the corresponding...following steps: Do a single or double digest to be sure that the plasmid is the correct expected size and...acknowledged in hundreds of publications including Cell, Nature, and other journals. Paolo Colombi is currently... -
Viral Vectors 101: An Introduction to AAV
TypeBlog PostPublishedMay 30, 2023, 1:15 p.m....systemically administered in an organism. These features make AAV ideal for human therapies and in vivo..., many of which are available in rAAVs! Figure 1: Diagram of AAV components Cargo capacity...gene switches, and/or serotype can also be used to ensure specificity; this is particularly helpful when ...differentiated, non-dividing cells. Figure 2: DNA rescue and transduction of a conventional...uncoating in the target cell nucleus, this DNA structure can readily fold into transcriptionally active...double-stranded form through intra-molecular annealing. Figure and legend from Okada, 2013. scAAV If your gene...As a starting point, we recommend checking the literature and our AAV data hub to see if others have used... -
Antibodies 101: Selecting the Right Antibody
TypeBlog PostPublishedMarch 15, 2022, 1:15 p.m....well for applications that denature a protein to its primary (linear) structure using heat or acid, for ...Picture this: you’ve been assigned an exciting new project aimed at understanding how a critical cellular...the application-specific data provided on the manufacturer’s website. Because of how they are developed,...Take a look at the publications cited on the manufacturer’s website, which may contain more detailed data...in a single experiment then you will need to make sure that you can easily distinguish each different type...epitope target, source, and other information to make sure you’re not trying to choose between the same antibody... partners by mass spectrometry (Uhlen, 2016). Be sure that the antibody you choose has been validated ... -
Celebrating 15 Years of Scientific Sharing
TypeBlog PostPublishedJan. 14, 2019, 2:08 p.m.... achieved and the opportunities we have for the future. At Addgene we will never ever stop planning, perfecting...re meeting the challenges of DNA distribution to ensure scientists everywhere can access plasmids despite... team. We show up in both science journals like Nature and in mainstream media like The Atlantic. We’ve...working in lock step to maximize the services and features we can offer online for Addgenies and to serve...Addgenies strong. The Addgene Board of Directors is mature and poised to support our growth. Thanks to Joanna...thinking, and openness to team work. Company culture Once in awhile I worry a very little bit about...Best Place to Work with a positive, supportive culture, we are attractive to the local employee pool and... -
5 Steps to Writing Grant Proposals for Grad Students
TypeBlog PostPublishedFeb. 11, 2014, 12:13 p.m....review the literature and offer educated insight into what the desirable features of future technologies...communication skills, Elucidates the big picture and future directions of the project, Outlines a detailed... proposed work. Thus, every single sentence and figure must be clear, concise, meaningful, and supportive...months to give feedback on the application. Make sure to invest time and consider the reviewer's comments...grant writing comes with continual practice and exposure, here are some additional links that offer more...first grant proposal" by Soapbox Science Editor in Nature Blogs "How to write a research project grant application... -
CRISPR 101: Targeting Non-Coding RNAs with CRISPR/Cas9
TypeBlog PostPublishedNov. 14, 2024, 2:15 p.m....transcription, and splicing. Nature. 2016;539(7629):452. doi:10.1038/nature20149 Jiang Q, Meng X, Meng L, ...this may still unexpectedly affect nearby genes (Figure 1A-C). Find CRISPR plasmids at Addgene! Knock-in...will terminate transcription of most of the ncRNA (Figure 1D). While this method can be time-consuming, it...-target effects (Zibitt et al. 2021). Figure 1: A schematic illustration of CRISPR/Cas9-based...insertion of a synthetic polyadenylation (spA) signal. Figure and caption adapted from Hazan and Bester 2021....nearby genes and interact with specific chromatin structures or other local genomic elements (Hazan and Bester...CRISPR/Cas9-targeting of long non-coding RNA genes. Nature biotechnology. 2020;38(5):573. doi:10.1038/s41587... -
Antibodies 101: Stripping and Reprobing Western Blots
TypeBlog PostPublishedAug. 1, 2024, 1:15 p.m....evenly re-absorb aqueous buffer. Be sure to check the manufacturer’s instructions for the membrane you ...companies including ThermoFisher, Millipore Sigma, and Azure Biosystems offer pre-made stripping solutions or...solution and agitate for 5–30 minutes at room temperature. The incubation time will depend on your blot... stripping solution uses heat and detergent to denature and remove antibodies but is also likely to remove...also volatile and toxic (and smells bad!), so be sure to use it under a fume hood. Rinse the membrane...Assess the stripping Now it’s time to check to make sure the antibodies from your first round of detection...remains, you may want to repeat your stripping procedure, perhaps using longer incubation times, or try... -
Tips for Technical Support Calls
TypeBlog PostPublishedSept. 20, 2016, 2:39 p.m....consider before you commit: Double check to make sure the answer to your question didn’t come with your...means their original articles are your best bet to figure out how to use a particular reagent. We also have... improve your customer service experience: Make sure you’re contacting the correct company. This might...possible about the issue. For Addgene support, make sure to mention the plasmid catalog number, or at least...experts on every technique or experiment, but a picture of a gel or a sequencing trace file can go a long...long way in helping us understand the nature of your concern. Please be courteous. We understand how... support that I miss is that you’re never really sure what you’re going to get. There are questions that... -
Visualizing Your Plasmid Request Data
TypeBlog PostPublishedApril 11, 2023, 4 p.m....excited to see that Addgene has recently added a new feature for depositing labs to its website, allowing depositors...aggregated data. Plotting the data in interactive figures is a great step up, but I needed a way to show ...Addgene Plasmid Request Data Dashboard! Figure 1: The Addgene Requests Dashboard The dashboard...following items (Fig. 1): A row with four of the key figures Number of requests per reagents (as a horizontal...request data to the dashboard (Fig. 2). Figure 2: Insert showing the upload function for the Addgene...depositor and the date of downloading the CSV is shown (figure 2) and the dashboard displays the data in a standardized...organization(s) is not shown on the dashboard. Key features of dashboards are (i) easy access to important... -
Finding Your Science Policy Path
TypeBlog PostPublishedSept. 22, 2020, 1:15 p.m....others (see more in Figure 2). Starting early demonstrates not only your interest to future positions, but ... Individuals that work in science policy are a treasure--they love the work and the mission so much that... Luckily, there are many mechanisms in place to ensure your effective networking---and they’re more accessible...of science with a large set of other societal pressures including economic, cultural, and political. Science...relations, advocacy, and writing legislation (see Figure 1 for career map). The beautiful part of science...more about science policy as a profession. Figure 1: Science policy can lead you to many paths! For...sabbatical year or transition away from academia as tenured professors. There’s never a bad time to start your... -
The Addgene Antibody Data Hub
TypeBlog PostPublishedSept. 12, 2024, 1:15 p.m....the antibody performed. Figure 5: Results for the data report shown in Figure 3. The antibody is rated... how an antibody performed in an experiment. Figure 1: The front page of the Addgene Data Hub. Antibody... whether the results include knockout data. Figure 2: Search options in the Antibody Data Hub. Data...find that information in the report as well. Figure 3: An antibody data report, showing the results...description of the experimental conditions. Figure 4: A screenshot of the materials and methods section...section for the data report shown in Figure 3. At the end of the report, you’ll find a results section... To submit, simply click "Contribute Data." Figure 6: Contributing data to the Data Hub is easy! ... -
The Scientific Conference Poster Session: Tips for Success
TypeBlog PostPublishedAug. 29, 2019, 12:50 p.m....papers or “hot topics,” so be sure to be well-read on the recent literature in your field. Take your time...about to present your first scientific poster, but unsure how best to prepare. Maybe you’re a presentation...attending, your preparation and day-of game plan will ensure you have the most beneficial scientific conference.... For example, when thinking about discussing a figure, jot down the question you were trying to answer... to say and provide your audience with a clear picture of your work. Practice, practice, practice: Rehearse.... Finding your poster board well ahead of time ensures you aren’t scrambling to tack up your poster seconds... the poster, as opposed to engaging with you? Be sure to use your poster as a visual aid, not a crutch...