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  1. Addgene-Seeding Labs plasmid grant round 3: microbial drug resistance and biofilms, antidiabetic plant components, visualizing brain injury - and a glowing robot fish

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    Blog Post
    ... Otito Izu is a PhD Candidate working under the supervision of Akindele Oluwatosin Adeyi at the University...University of Ibadan, Nigeriaunder supervision of Dr. Akindele Oluwatosin Adeyi, University of Ibadan, Nigeria...research related to space medicine. Now he aims on tracking pollution in rivers by employing synthetic biology...be able to support diverse groups of scientists working in a wide range of research fields and geographical...
  2. "Build Your Plasmid": The Game - Play to Teach

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    Blog Post
    ...Professor at University of Barcelona. The challenge: Making courses on genetic manipulation more hands on While...meeting with some other professors, I went home thinking about how to solve this problem. I didn’t want...your strategy. I realized that similar types of thinking go into developing gene therapy strategies for...noting any gaps in their knowledge. Practically speaking the format I decided on was to have two teams ...skills like team building, collaborative critical thinking, and communication among others. They additionally...
  3. FPbase: A new community-editable fluorescent protein database

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    Blog Post
    ...much of the primary data one might require when making such a decision remains spread across literature...With thousands of scientists routinely using and thinking about FPs, it would be great to have a centralized...building advanced multi-parametric search queries.  Looking for a bright, monomeric FP published in the last...suggestions. If you’re not sure exactly what you’re looking for, the Table of Fluorescent Protein Properties...that set of proteins.  For computer-savvy users looking to incorporate FPbase data in their own projects...
  4. Crowdfight, a Platform to Boost Scientific Collaboration During COVID-19 and Beyond

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    Blog Post
    ...culture towards a more generous and rewarding way of making science.   How we started Crowdfight-COVID19...as a collaborative platform to help researchers working on COVID-19. Its origins trace back to March 2020...-19, an online platform to connect researchers working on COVID-19 with scientists of all areas who volunteered...idea behind the platform was simple: any person working on COVID-19 could file a request on our website...We are a community of researchers who believe in making science less of an individual effort and more of...
  5. Visualizing Translation at the Single Molecule Level

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    Blog Post
    ...monitoring the first round of translation or by tracking the translation of a single mRNA over time. Both...translation. Single molecule continuous translation tracking TRICK allows you to see the first round of translation...translation, but it doesn’t allow continued tracking of a given mRNA because NLS-PCP-GFP is removed by ...first ribosome that translates the mRNA. To permit tracking over time, Yan et al. devised a system that labels...keeps it in a single field of view for constant tracking. You can then directly monitor multiple rounds...
  6. What Good Citizenship Can Do for Reproducibility in Science

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    Blog Post
    ...important part of being a “good scientific citizen” and making sure you do what you can to help your fellow researchers...consistent way so other researchers can use them, to making sure that methods and protocols are available and... can be published. To a scientist, all of this checking and formatting can feel like yet another hurdle...for being transparent in their communication or making their data and reagents readily available. What...confident that everyone’s enthusiasm for actually making it happen would increase dramatically. I’m not ...
  7. Selecting Your Plasmid Purification Kit

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    Blog Post
    ...something is working for you, then there is no reason to change it. But what if a kit isn’t working for you...ethanol, or endotoxins. The most common method for checking both quantity and quality of your plasmid prep...spectrophotometry. Achieving the highest-quality DNA requires taking care at each step, but some purification kits make...secret that labs produce a lot of waste. If you are looking to reduce your lab’s environmental impact, check... and yes those purification columns we’ve been talking about!) are a significant source of waste in laboratories...
  8. How to Deposit Your Plasmids with Addgene

    Type
    Blog Post
    ... account, you can begin the deposit process by clicking "Submit Online Deposit" (for online submission... menu: Figure 1: Begin the deposit process by clicking "Submit Online Deposit" or "Download Deposit Spreadsheet...  Figure 4: Add plasmids to the article by clicking “Add Plasmid to this Article” (left) and entering...button and jump around to the different pages by clicking on the appropriate boxes at the top of the page...the plasmid to copy from the drop-down menu and clicking the white "Copy" button. *Note: The "Copy" button...Submit Deposit" button to complete the data entry. Clicking the "Submit Deposit" button will allow you to ...
  9. Early Career Researcher Toolbox: Social Media for Scientists

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    Blog Post
    ... in Your Working Day" video from Science Media Savvy At this point, you might be be thinking social media...first blog post out of boredom. I had just started working in a new lab and I was the only person in the lab...first post so I signed up to write a second. While working on this post, I was offered and accepted a position...on the Addgene blog Read about online social networking for scientists Learn why scientists should be...
  10. From Lab Bench to Leadership: Transitioning into Management as a PhD

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    Blog Post
    ...responsibilities. After my second postdoc, I realized that working in the wet lab wasn’t the right path for me. I ...style of a new, junior team member who had values, working styles, and skills that were quite different from...from them. I started at my previous company by taking on responsibilities such as mentoring a junior ...Beyond Academia: Stories and Strategies for PhDs Making the Leap to Industry." He successfully transitioned...Resources  Resources for aspiring managers Books: The Making of a Manager by Julie Zhuo Radical Candor by Kim...
  11. Plasmids 101: Inducible Promoters

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    Blog Post
    ...protein AraC binds O and I1 sites upstream of pBad, blocking transcription. The addition of arabinose causes... cAMP and represses pBad, decreasing promoter leakiness.   Other examples of chemically induced promoters... be compatible with your experimental system. Leakiness If you’re expressing a gene that may be toxic,...inducible promoters are also known for their low leakiness. Inducibility level How much do you want to induce... inducible promoters can vary a lot. If you’re looking for high inducibility, the Tet-On system may be...
  12. Antibodies 101: Buffers, Storage, and Conjugates

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    Blog Post
    ...can leave them at room temperature when you’re working with them, but avoid freeze-thaw cycles, as they...bind to amine groups present in some conjugates, blocking the conjugation reaction. Many commercial antibodies...for your ELISA and Western, I’d highly recommend sticking with what works.  To balance this out, however...in your machine before you make your purchase. Picking a single fluorophore for an experiment is easy,...accurate read and reduce needed compensation.    Working with antibodies can get a bit complicated, but ...
  13. Sharing is Caring: But How to Distribute Open Hardware?

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    Blog Post
    ...toolbox openUC2 are trying to overcome this hurdle by taking these limitations into account in the development...software (left) vs. open hardware (right).    Taking this all together, the essential ingredients of...newly developed techniques are the key enablers for making assemblies open-source. The sharing of digital ...produced quickly with the printer, act here as a kind of "teleporter," since a part developed in Germany...way of communicating in science brings stronger networking across laboratory and disciplinary boundaries...
  14. How Addgene Creates Content

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    Blog Post
    ...channel just for asking people if they can write or review things and another for asking specific science...depositors and customers are asking. So how can you get involved? Keep asking your burning questions and...review to catch those pesky typos, and hit publish. Making content is an iterative process. We are constantly...
  15. Channelrhodopsins with improved light sensitivity for minimally-invasive optogenetics

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    Blog Post
    ... the mice to increase their nose-poking behavior. Strong nose-poking behavior is exactly what was seen...a channelrhodopsin, such as cell localization, kinetics, and spectral properties, must be optimized simultaneously...photocurrent strength, wavelength sensitivity, and off-kinetics.   This model was then used to predict which channelrhodopsins...photocurrent amplitude, wavelength sensitivity, and off-kinetics correlated with the model’s predictions.  Three...
  16. RNA Extraction Without A Kit

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    Blog Post
    ...on everything, including your skin. Here are some general tips for working with RNA, even if you are using...basics of isolating RNA without a kit. Tips for working with RNA (whether you are using a kit or not) Although..., 2006).  View our protocol for RNA extraction! Making the simple protocol even more foolproof (still ...still without a kit!) As mentioned above, working with RNA requires keeping your samples cold until homogenization...
  17. The time and cost required to make a plasmid

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    Blog Post
    ...digest/ligate methods and picking the right one can save you time if you’re working at speed or money if budget...you can explore to maximize the use of both when making plasmids. Use repositories Repositories such as...free. Plasmids ship out 2-3 days after ordering, making it a toss-up as to whether it’s faster to order...experiment. Make your work “high-throughput” Instead of making one plasmid at a time, try constructing several...
  18. CRISPR 101: Cas9 vs. The Other Cas(s)

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    Blog Post
    ... unique cutting features and evolved PAM sites, making Cas9 an adaptable choice for many genome engineering...same mechanism as traditional spCas9. If you are looking to modify RNA, Cas9 just won’t cut it – literally...editing. Fast facts – PAM requirement: protospacer flanking sequence – A, U, or C. Best for: targeting single...use. Fast facts – PAM requirement: protospacer flanking sequence – none. Best for: targeting single stranded...single base pair edits, these enzymes are worth taking a look at. Choosing the right Cas can really put...
  19. Developing Lab Management Software for Biology

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    Blog Post
    ... it’s monumentally difficult without software. Tracking lab procedures also gives you access to hard data...repeated. We also now have a good interface to begin tracking analytics data for lab procedures. With this data...are distinct from their contents. You should be tracking both, you should easily be able to move contents...less time getting up and running and more time working on interesting biotechnology code.There are valid...reasons to choose a non-Python stack, but if you're working with scientists it's unlikely they will have exposure...
  20. The Future of Research Symposium Boston 2015

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    Blog Post
    ... There are currently 17 sites around the USA undertaking BEST programs, including local schools in Boston...contributing to the NIH’s Biomedical Research Workforce Working Group. Along with her participation in this panel...Pubmed Central PMCID: PMC3787295. 6. Marder, Eve. "Looking out for future scientists." eLife 3 (2014): e04901...PMC4185419. 7. NIH 2012. Biomedical research workforce working group report. http://acd.od.nih.gov/biomedical_...in Science in Our Career Posts! Get Advice on Networking at Scientific Meetings from Addgene Executive...
  21. Prime Editing: Adding Precision and Flexibility to CRISPR Editing

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    Blog Post
    ...edit and homology to the genomic DNA locus. After nicking by Cas9, the homologous pegRNA sequence hybridizes..., as it can edit near to or far from PAM sites, making it less constrained by PAM availability than other...previously used when they developed base editing. By nicking the non-edited strand, they can cause the cell ...efficiencies 2-3-fold. However, the process of double nicking increases indel formation slightly. Designing the...the edited strand, as in the PE3b system, guides nicking of the unedited strand only after editing has occurred... ✓ ✓ ✓   ✓ Second sgRNA to guide nicking of the unedited strand Favor the edited strand...
  22. A Guide to Designing a Scientific Poster: Content, Layout, and More

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    Blog Post
    ...include a lot more. Font size: Making your poster readable While we are talking about font, remember this:...important to reflect on these impressions when thinking about crafting your own poster. What sets a scientific...that it will display your data most prominently, making it the star of your presentation. Portrait. This...presented some standard formatting guidelines for making a scientific poster, you shouldn’t hesitate to ...
  23. Viral Vectors 101: Virus Safety

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    Blog Post
    ...understand the risks of the specific virus you’re working with before you work with it. This will help you...required in BSL-1 and required in BSL-2 labs. When working with lentivirus, many organizations require or ...can quickly be removed and disposed of without risking exposure. You can request disposable gowns from...researchers choose to wear eye protection when they are working in a biosafety cabinet, even if it is not required...used for BSL-2 waste, and of course, if you are working with a substance in a biosafety cabinet, it needs...
  24. Gaining Leadership Skills Volunteering at a Professional Organization

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    Blog Post
    ...with the NPA where I gained invaluable experience working on and later chairing the Advocacy Committee. I... I’m now furthering my leadership experience working at the Association for Research in Vision and Ophthalmology...website. By volunteering, you can gain experience working with other people mostly online and on monthly ...resulted in actionable tasks and I made a point of sticking to deadlines and performing the work I agreed ...members changed, I had to learn how to develop new working relationships with these individuals who often ...
  25. Story of a SynBio Startup: RevBio's Epiphany (or Lack Thereof)

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    Blog Post
    ... months have in store for us. If you’re thinking about making the switch from academia to entrepreneurship...entrepreneurs. To make things more interesting, we are working in a new technical discipline that nobody understands...few jobs available. So, we spent a lot of time talking about what we could do to make ourselves marketable...marketable to the wider world and occasionally thinking about how we could bypass the establishment hiring...
  26. CrispyCrunch: High-throughput Design and Analysis of CRISPR+HDR Experiments

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    Blog Post
    ...a bench scientist would looking at a genome by eye. We came up with a ranking formula that does just this...In the rest of this article, we'll explain the thinking behind the tool, its key features, how it works...information from Crispresso plus a summary per batch. Ranking guides for HDR in-depth In other tools, guides ... engineer with the Chan Zuckerberg Initiative, working on assignment at the Biohub in San Francisco. Before...
  27. Degrading DNA with Cascade-Cas3

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    Blog Post
    ...broken down into different types, with Cas3 systems making up type I, the most abundant type in the Class ...consistent expression can still be a challenge. A multitasking Cas Most Cas effectors have one job — Cas see...away at both the non-target and target strands by making cuts every so often. Cutting both strands ensures...carried out much like the name implies — after nicking, Cas3 reels in the non-target strand of DNA much... for Cas3’s biggest trick. Cas3 likes to bluff, making you into think it’s more powerful than it is. The...
  28. Plasmids 101: 5 factors to help you choose the right cloning method

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    Blog Post
    ...You’ve spent days and weeks thinking of an amazing project. You’ve written your protocols, designed your...enzymes laying around the lab. If not, you’ll be looking at about $50-$60 per enzyme. Once you’ve got a ... of the restriction enzymes and overall you’re looking at a price of a few dollars per reaction for traditional...relatively new to the cloning game, however, or simply looking for a more streamlined approach, you might want...inserts: There are some cloning methods that make working with multiple inserts a breeze. Among them are ...
  29. Cloning Mammalian Cells with the Agarose Method

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    Blog Post
    ... they become confluent, taking both early- and late-arising clones, and making sure we always maintain...densities - adding to the expense of dilution cloning. Taking a page from bacterial cloning, we reasoned that...cloning rings. A picture of a 60 mm dish ready for picking is shown in figure 1.   Find Additional Useful ...firmer 2% agarose gel solution discussed above for picking. Many thanks to our guest blogger Iris Lindberg...
  30. Fluorescent Proteins 101: Visualizing Subcellular Structures & Organelles

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    ...Additional opportunities: Tracking intracellular pathogens In addition to tracking the location of proteins...University of Queensland) to observe membrane trafficking events in vivo. Locate your protein and see who...your images using subjective judgement by simply “looking at them”. Modern cameras and image analysis software...limited by the wavelength of light and, practically speaking, for common lab microscopes the detection limit...
  31. Plasmids 101: Knockout/Knock-In Plasmids

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    Blog Post
    ...alleles. Although CRISPR is very good at making knockouts, knocking in large sections of DNA can be more ...negative selection marker like the HSV thymidine kinase (HSV-tk) is included just outside one of the homology...” the resistance gene through the insertion of flanking LoxP sequences, you can remove the gene through...described in the preceding sections are simple ways of knocking out a non-essential gene of interest. However,...
  32. Addgene's Cancer Collection Pages Connect the Research Community

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    Blog Post
    ...Despite these grim statistics, new strategies for attacking cancer are yielding promising results in the laboratory...universally-dividing behavior of cancer cells, attacking the cellular machinery necessary for division....in the field of cancer research has focused on picking apart the complex cellular pathways involved in...found across all TCGA tumor samples is represented, making the results of any such study as clinically relevant...Addgene's cancer collection pages Scientists are making progress. New resources and technologies become...
  33. Drew Endy Introduces the Biobrick Public Agreement Plasmid Collection

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    ...co-founder of the BioBricks Foundation, an organization working to advance biological engineering openly so as ...thing at a time. What the BioBricks Foundation is seeking to accomplish is to make the engineering of biology...is in reference to George Boole who in 1854 was thinking about logic and analyzing human language and observed... be made freely available? Endy: When we start thinking about languages, humans have two types of languages...really plugging into the "stuff sector" if you will, making chemicals and materials, and we anticipate plugging...
  34. Deep Dive: qPCR

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    ...using the application correctly is imperative. Picking your chemistry  The two most common qPCR chemistries...thermodynamic and experimental parameters (neighbor stacking energies, loop entropy effects, cation concentrations...type (such as here, here, or here).  If you’re working with samples that have not been documented in the... plateau stage after exponential amplification, making it difficult to properly analyze your data.  To...experiment! The model assumes the amplification kinetics of the target and control genes are fairly equal... e-book on applying MIQE guidelines! With these kinds of resources, you will soon become a qPCR master...
  35. CRISPR/Cas9 FAQs Answered!

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    Blog Post
    ...to have 50–80 bp of homology arms on each side flanking the change you'd like to introduce. We use Ultramer...plasmid donor, with two homology arms on each side flanking your desired insertion or mutation. Each arm is...choosing target sites that span the knock-in gene. For making mutations, one good way is to mutate the PAM 'NGG...Matoba, S., Zhang, Y., & Zhang, F. (2013). Double nicking by RNA-Guided CRISPR CAS9 for enhanced genome editing...cells differing exclusively at two early onset Parkinson Point mutations. Cell, 146(2), 318–331. https:...
  36. Capturing a Moment with Marc Zimmer’s “The State of Science”

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    ...Science: What the Future Holds and the Scientists Making It Happen, is an exercise in restraint. The very... revelations. The issues he discusses have been making the rounds in journal editorials, science social...likely range from mildly surprising to genuinely shocking. Mainstream media outlets have addressed CRISPR...recontextualized some areas of science that I’ve been thinking about a long time. For the scientists who like... Eric J. Perkins...
  37. Negative Can Be Positive: Open AAV Data with Addgene

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    ...Working with AAV vectors in the brain and peripheral nervous system presents a special challenge to scientists...answered this question and used it to publish!  Working with AAV vectors in neuroscience Let’s consider...consider my first challenge when I started working with AAV vectors. I wanted to use an AAV vector to express...label neurons transduced by each serotype. Working at Addgene has shown me this was far from an isolated...Results from a survey conducted by Addgene in 2021 asking scientists what data would improve our AAV Data...
  38. Lentiviral Vector Uses and Overview

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    ...PMCID: PMC110254. 3. In-vivo RGB marking and multicolour single-cell tracking in the adult brain. Gomez-Nicola...pre-clinical/early clinical stages. Researchers are also working to make lentiviral vectors safer. Since lentiviral...Winemiller A, Andre K, Stein D, Blick G, Greenberg RN, Kinder C, Zolopa A, Binder-Scholl G, Tebas P, June CH,...therapy approaches to sickle cell disease. Hoban MD, Orkin SH, Bauer DE. Blood. 2016 Feb 18;127(7):839-48. ...
  39. Plasmids 101: Protein tags

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    Blog Post
    ... a molecular leash for your protein. If you’re working with an uncharacterized protein, or a protein for...for antibody-based purification; has inherent enterokinase cleavage site GST Large Protein 26 Purification...context of a cell. FLAG tags can be cleaved by enterokinase, which recognizes DDDDK^X, cleaving after the...rounds of purification, with the cleavage reaction taking place between column runs. The TEV protease itself... Eric J. Perkins...
  40. Interview: Hodaka Fujii on enChIP, New CRISPR Tools, and More

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    ...Fujii: In the last several years, my lab has been working on development of technologies for biochemical ... molecules even enables us to skip the step of knocking-in. In 2013, we showed that isolation of a single...in the near future. Addgene: Has your lab been working on any other technologies that you would like to...that involvement in the scientific community is a kind of duty for researchers when considering the peer-review... in my lab then when they become independent. Looking for enChIP Plasmids? All Hodaka Fujii lab plasmids...
  41. Negotiating Work and Life: How to Find the Joy

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    ...that end, here are my personal 10 commandments for making it all work – most of the time. #1 Thou shalt find...happy at work, you will find balance. If you enjoy working and come home happy in the evening, your friends...of support to help with elder care, carpooling, cooking on nights when you can’t get home, babysitting ... really hard not to do this. For a family with working parent(s), the sooner your kids can make a decent...(except I am glad she never told me how much hitchhiking she did in Israel until after the fact). #9 Thou...
  42. Plasmids 101: Shuttle Vectors

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    ...unrelated host species, even from completely different kingdoms of life. That’s convenient for researchers who...example, puromycin inhibits protein synthesis across kingdoms by targeting a shared aspect of ribosomal function... Zeocin, a DNA intercalator, which affects all kingdoms of life. Of course, that includes you. Besides...plasmids are those that can replicate in multiple kinds of bacteria, including many natural plasmids. By...vectors can help you hop, skip, and jump between all kingdoms of life. A bit of careful design will help make...
  43. Twenty Years of Sharing: Expanding Accessibility

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    ...amazing 111 countries (Figure 1)! But while we were making the materials available to many countries, import...stays in transit for a certain number of days and tracking average transit time for each courier we contracted... to increase accessibility to our repository by making it easier to find materials and learn about them...information are truly accessible, not just available.   Looking forward Over the past twenty years, Addgene has... accessible to scientists across the globe?” By asking this question, and by listening to the answers ...
  44. Neuronal labeling with Spaghetti Monster

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    Blog Post
    ... used for protein localization, isolation, and tracking experiments (Rizzo, 2009). With many different...aggregation and cross-react with anti-GFP antibodies, making them difficult to use in a multicolor/multichannel...fluorophore-labeled antibodies with very specific reactivities making them more useful for multicolor imaging experiments...Davidson, and David W. Piston. "Fluorescent protein tracking and detection: fluorescent protein structure and...Blog Rabies and neuronal tracing Retrograde AAV: Making the journey from axon to nucleus AAVs CREATEd for...
  45. Hot Plasmids - November 2023

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    ...cargo Optogenetic proximity labeling and TransitID Tracking cellular history with protein ticker tape Teaching...effective and with no additional off-target edits, making them ideal for size-limited AAV delivery. PE6c ..., et al. (2023). Dynamic mapping of proteome trafficking within and between living cells by TransitID.... bioRxiv – doi: 10.1101/2023.02.07.527548.    Tracking cellular history with protein ticker tape By Brian...call for thin sections of tissue, the Wu Lab is working to create a three-dimensional atlas of protein ...
  46. Plasmids 101: The Promoter Region – Let's Go!

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    ...make sure your plasmid has a promoter suited to working in your host organism. Because transcription machinery... and there is very little crossover. Generally speaking, promoters in bacteria are less diverse and complex...elements, insulators, and silencers; however, some "leakiness" of transcription may occur. This is normally ...expression mRNA Mammalian promoter from phosphoglycerate kinase gene.  Constitutive  Widespread expression, but...optogenetics mRNA  Ca2+/calmodulin-dependent protein kinase II promoter Specific Used for neuronal/CNS expression...
  47. Welcome to the Lab Bench! New Protocol Series Helps Trainees Get Started in the Lab

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    Blog Post
    ...New to the lab? Or looking for ways to supplement your teaching? You’re in luck! Addgene’s Intro to the... We had many protocols for plasmid cloning and working with viral vectors, but what we lacked were videos...guiding completely new members of the lab or those taking introductory lab courses. The Intro to the Lab...science techniques. It’s also great for teachers looking to supplement their curriculum. From the details...
  48. New Optogenetic Tools for Cytoskeleton and Membrane Control

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    Blog Post
    ...the specific Rho GTPase/GEF pair (Ridley, 2016). Taking control of a RhoGEF is a great way to stimulate..., 2023). Rab family proteins support vesicle trafficking and organelle dynamics, but with dozens of family...Optogenetic inhibition of Rab protein vesicle trafficking via CRY2-CIB1 clustering. Image adapted from ...chemical composition of the membrane itself? The Baskin Lab previously published OptoPLD, an optogenetically....2015.08.005.  Tei, R., Bagde, S. R., Fromme, J. C., & Baskin, J. M. (2023). Activity-based directed evolution...
  49. An Integrin Antibody Toolkit from IPI

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    ...express multiple integrins that bind the same ligand, making it difficult to dissect the function of one integrin...antibody is extremely rare. While many function-blocking integrin antibodies exist, almost all bind outside...Springer lab, reported the binding specificity, kinetics, and affinity of these 11 newly discovered synthetic...to the Springer lab, a highly specific function-blocking antibody, αVβ3.13, may also block function of ...important integrin on these cells. What’s next IPI is working with Addgene to accelerate biomedical research ...
  50. Enabling high school research at the Journal of Emerging Investigators

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    Blog Post
    ...protein required for cytokinesis in JEI. Developing an interested in biology, working in a lab for the first...(the importance of word of mouth and personal networking!) and the manuscript review/submission process... first time, and thinking about the future. Many thanks to our guests,Olivia Ho-Shing, Mark Springel,...
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