... developed during the COVID-19 pandemic. Our mission to accelerate research through sharing reagents has...2021.03.10.434840 Tsang J, LaManna CM (2020) Open Sharing During COVID-19: CRISPR-Based Detection Tools. The...detect SARS-CoV-2 nucleic acids. Mammalian targets During the 2003 SARS outbreak, Hyeryun Choe’s lab identified...-CoV-2 entry into human lung epithelial cells (Biering et al., 2021). In their search, they found mucins...inhibitors of SARS-CoV-2 entry and become upregulated during viral infection.  Expression of SARS-CoV-2 viral...academic and industry scientists through plasmid sharing and more. As you’ve read above, some of these plasmids....     References and resources References Biering SB,  Sarnik SA, Wang E, Zengel JR, Sathyan V, Nguyenla...

...by Benoit Giquel. CRISPR has taken the genome engineering world by storm owing to its ease of use and utility...Waltz, 2016), the potential of CRISPR for genome engineering in plants is also being realized. There are a...approaches, these techniques are cumbersome, often requiring several rounds of selection to isolate plants ...plants with the phenotype of interest. Genome engineering, on the other hand, allows for targeted modification...will present a general overview of plant genome engineering using CRISPR, highlight the specific modifications...plants, and outline the various plant genome engineering tools that are available to academic researchers... Addgene. CRISPR components for plant genome engineering CRISPR can be used to knockout, activate or repress...

...important role in scientific exchange. In addition to ensuring affordable access to materials, repositories provide...since 1996.       More Blog Posts about Scientific Sharing: The 10 Most Distributed Plasmid Technologies ...10 Years Data Freedom: The Expansion of Data Sharing in Research Publications ...

...career path and why he chose to go into genome engineering. Chapter 1: Early research on CRISPR and what...with new project ideas, applying for grants, and mentoring. Chapter 5: Wei Leong Chew's lab management style... 6: Projects in Wei Leong Chew's lab. Genome engineering, gene delivery methods, AAVs, and more. Chapter... genome engineering...

...large scale cell mobility are essential processes during embryonic development. These are required for a...development of cell sorters together with the engineering of fluorescently labelled antibodies made it ...)? How do blood monocytes patrol blood vessels during immune responses (Auffray et al. 2007, Finsterbusch...relatively little tissue absorption, weak tissue scattering and small amounts of tissue auto-fluorescence...understanding the key roles of immune cells in triggering an immune response. New fluorescent proteins ...Dynamics of neutrophil migration in lymph nodes during infection." Immunity 29.3 (2008): 487-496. PubMed...Jackson G., et al. "Macrophage and T cell dynamics during the development and disintegration of mycobacterial...

...raise salaries prior to the injunction are still honoring these plans. We are now collecting data on the...who had a contract - negotiated by their union - securing higher salaries. This decision will carry over...geographical regions When we originally started gathering this information, many assumed that those living...Read Gary's Previous Article: Changing Labor Laws Bring Increased Postdoc Wages Read about the Advocating...

... to academic scientists. Expanding plasmid sharing during COVID-19 Realizing this limitation, we worked...members of the scientific community support open sharing. But, one critical part was missing when the COVID...setting.  In light of the need for basic research during the current pandemic, universities are showing ...cells. We teamed up with Ginkgo Bioworks to quickly bring these plasmids to the scientific community - both...available to industry scientists To make plasmid sharing more open for academic and industry scientists,...the past months, we’re excited to see our open sharing impact grow beyond COVID-19. But, we can’t do this...office to discuss the possibilities for expanded sharing. Is there a plasmid you're hoping will be come ...

...contributions from depositors around the globe, we’re now sharing over 100,000 reagents including plasmids, ready-to-use...

...these extreme environments. Gearing up for sample collection at the hot springs My husband, Paul Blainey,...all of the remote areas, including one hot spring requiring an overnight hike in. Navigating challenges...the hot spring. Hiking all day and processing samples by night was fun but also very tiring. It feels...Lab at Stanford University became interested in exploring biological dark matter – large tracts of the microbial... these microbes. From 16S rRNA sequencing, hot springs were known to be diversity hotspots containing ...aquaticus bacterium – was discovered at Octopus Hot Spring at YNP, and since then, many researchers have searched...temperatures and unstable ground around these hot springs make them extremely dangerous, we used telescoping...

...Addgene Blog Mentoring for Scientists Series. I have been thinking a lot about Mentoring for over 10 years...group mentoring format could work wonders.  I have now seen the formation of hundreds of mentoring groups...also serve as a great first-time mentoring experience. They make mentoring less mysterious and train the ...useful for future mentoring relationships.   If you would like to start a group mentoring program at your...interested peers and form your own peer mentoring circle. In a Peer Mentoring Group all participants act as both...and what you want to cover during the year. Suggest that each person bring 3 topics and write them on ...involved in Peer Mentoring relationships? Comment below with ideas from your own mentoring group or with ...

...to learn about her mentoring style. Top 7 questions scientists ask me about mentoring In this blog series...the other six mentoring questions in future posts. If you have a question about mentoring that you want... enough?  For many years I’ve been organizing mentoring programs for scientists and doing training to ...fulfilling, productive relationships. I will be sharing what I’ve learned along the way in this ongoing... to the top questions scientists ask me about mentoring. Here they are: What makes a good Mentor? How...? What is this I hear about “group” or “peer” mentoring? How do I ask someone to be my Mentor? I have ...certainly realized the importance of seeking out mentoring as an important aspect to their ongoing success...

...KTRs are not well suited for monitoring kinase activity that occurs during nuclear envelope breakdown (...important for regulating signal transduction and measuring kinase activity at the single-cell level can aid...signaling activity and cell phenotype. One method for monitoring live single-cell kinase activity is FRET, but...Advantages of KTRs When compared to FRET reporters for measuring kinase activity, KTRs have two key advantages....

...bacterial CRISPR engineering One common way of accomplishing bacterial genome engineering is done with recombineering...combines CRISPR and recombineering-based MAGE (Multiplex Automated Genome Engineering) technology (Ronda...combines recombineering with CRISPR to create a system for scarless, iterative genome engineering (Jiang...promoters in Pseudomonas syringae and Klebsiella oxytoca. Microbiome engineering Scientists have begun to...discovered in bacteria, most CRISPR-based genome engineering has taken place in other organisms. In many bacteria...quirks change the way CRISPR-mediated genome engineering functions in bacteria, but have no fear - plasmids... recombineering, a technique that utilizes phage recombination machinery to promote homologous recombination...

...multiplex genome engineering. Image from Sakuma at al. (2015). Multiplex genome engineering demonstrated...system for CRISPR/Cas9-mediated multiplex genome engineering is now available thanks to researchers at Japan...making an all-in-one vector for multiplex genome engineering.” Now, they’ve filled that gap. As Sakuma and...efficient viral vector-mediated multiplex genome engineering. The system now provides an efficient targeting...Sakuma’s protocol. Reference: Multiplex genome engineering in human cells using all-in-one CRISPR/Cas9 vector... multiplex genome engineering...

...database and ordering platform soon to be released. For individuals interested in ordering from ABRC for...Learn how BioBuilder can bring Synthetic Biology into the classroom Engineering the Plant Genome Using ...to us at [email protected] for help setting up an ordering account. Using Arabidopsis in the classroom has...CRISPR/Cas9 pSiM24: Simplifying Plant Genetic Engineering Resources on Addgene.org Find Plant Vectors...

...platform for sharing, annotating, and storing life science protocols. Since we began sharing this protocol...Developing Lab Management Software    Organizing and sharing your updated protocols So that’s great, but what...Plasmids 101 blog series. Want to learn more about sharing your plasmids via Addgene? Check out all the benefits...feature further simplifies lab work if you like to bring your phone in the lab, though, honestly, this still...non-bench work – but is there a sterile, clean way to bring your phone into the lab? Maybe having a dedicated... protocols listed on OWW, including those for preparing your own competent cells. If you’re familiar with...exchange currently boasts over 2,500 protocols covering a variety of bio and biomedical topics. Protocol...

...using glass cloning rings to isolate colonies of antibiotic-resistant cells; during the time required to... to grease, place and fill a dozen cloning rings, the remainder of the colonies on the plate dried out...out and died. The alternative to cloning rings, dilution cloning into 96-well plates, is extremely time...minutes- a far more rapid pace than using cloning rings. A picture of a 60 mm dish ready for picking is ...differ grossly from those isolated using cloning rings in terms of growth rates, morphology or other physiological...

...bad news. The bad news is that some of tubes’ o-rings are, in fact, permeable to CO2 at low temperatures...What we found Not all tubes (or rather, not all O-rings) were permeable to CO2 at low temperatures. We tested...Conclusions and handling recommendations Not all O-rings function optimally at -80°C. For samples that are...or not the dry ice we use to keep viruses frozen during shipment was having an impact on the samples. We... Order Ready to Use Viral Preps from Addgene Measuring the pH of our samples: What we did To measure ...CRIPSR User (By a 1st Time CRISPR User) Tips for Titering Your Lentivirus Preps Additional Resources on...

...: they require switching between a two-ring purine and one-ring pyrimidine base. This is a far more complex... the side groups of DNA bases and leave the core ring structure intact, allowing them to make 4 out of...backbone, making them efficient and precise tools for altering DNA sequences. These first base editors were both...researchers in the Keith Joung lab, who were engineering an ABE to reduce its tendency to make off-target...sites. To explore this further, the team started engineering a CBE to be worse at its original job. They ...glycosylase inhibitor (UGI) components, which inhibit BER during cytosine-to-uracil editing, and instead introduced...GBE editing but not CBE editing, maybe because repairing a GBE-derived abasic site requires more cellular...

...underside of the “umbrella” of A. victoria there is a ring that glows with a faint green light. To investigate...

...accessible and easier to share. As we prepare to ring in the New Year, let's make a toast to open science...those of us at Addgene believe in the power of sharing, and especially in keeping the products of science...

...give to your questions. If one of their answers rings true for you, go ahead and give a similar answer... to the hiring manager and others, if you have their emails. You can use notes you took during your interviews...Fielding questions can be hard during this talk since you’ll be sharing your screen and won’t be able ...2016 and then working four more years in academia. During my job hunt, I quickly noticed one of the biggest... and/or small groups, usually starting with the hiring manager. These interviews lasted for 30 minute ...a list of who will be interviewing you, ask the hiring manager or recruiter for this information. I found...presentation to your audience Start by asking the hiring manager or recruiter what presentation format they...

...a protein based on the affinity of the imidazole ring of histidine for metal ions such as Ni2+ or Co2+...the polyhistidine tag? The polyhistidine tag is a string of 6 or more histidine residues expressed in frame...step to remove the tag in order to prevent it interfering with protein function. In contrast, the polyhistidine...using these more selective substrates along with stringent wash conditions.   Find polyhistidine tag plasmids...Sometimes a tag placed on one end will be blocked during protein folding, in which case you should try tagging...neither placement works, consider purifying under denaturing conditions. Unlike other affinity purification...). In some cases, proteins can be bound under denaturing conditions to expose the tag and then refolded...

... position. Moving one position over in the sugar ring, a popular stabilizing modification is 3’ phosphorothioate...

...Stephen, Andrew G., et al. "Dragging ras back in the ring." Cancer cell 25.3 (2014): 272-281. PubMed PMID:...computer modeling have enabled researchers to begin discovering and designing molecules that are showing some...

...discovered that incorporation of four-membered azetidine rings could substantially enhance the brightness and photostability...

...actually has chimes outside of our break room. A ring of the chimes means something fun is going on and...in a unique environment where, in addition to furthering our mission by hard work, we genuinely enjoy ... that you don’t work for Addgene (though we are hiring!) but to encourage everyone to make an effort to... the power of simple word of mouth - be sure to bring up your event in casual conversation to get people...

...to help students during an interview. When they were instructed to use the three-ring binder as a starting...transferable skills learned while working in the lab and offering advice on areas where you might need some help...creativity is absolutely a transferable skill developed during PhD training. Creativity is required to develop...Whether it’s finding the source of contamination or figuring out why the lab is so cold, we’ve all had to think... need ahead of time and mute your notifications during. (Boyes, 2019) If you’ve taken the time to do this... assure that all participants feel comfortable sharing their ideas? And how do we give people the opportunity... The purpose of this form was to encourage the sharing of ideas, not to give people more work. Identify...

...camera lens without having to physically rotate the rings around the lens. It allowed me to focus on the subject...constraints given. And there were many constraints during the COVID-19 pandemic. The Pandemic Pivot Back...Even Blugene is practicing social distancing and wearing their mask. But while the camera is here, the operator...chair not watching their actors live on set, but peering at a 7-inch wireless monitor. Wireless video technology...control, and extra hand sanitizer.    Cycle 4: Venturing into the Lab Design: The lab is the beating heart...heart of Addgene. It’s why we exist, storing plasmids, and distributing them to labs that need them. The...projects lined up, some that started production during the middle of lockdown, and some that will spin...

...spent hours in the biosafety cabinet with cloning rings or following labor-intensive limiting dilution protocols...is used for a variety of applications such as measuring target protein expression levels, assessing post-translational...uses 2 or 3 different fluorescent colors each measuring a different target. Flow cytometers however are...methanol, fix and permeabilize in a single step by denaturing proteins and dissolving lipids including those...specialized components that can divert a subset of cells. During FACS, the sample stream oscillates to generate ...be targeted with specific fluorescent antibodies during the flow cytometry experiment and will appear as...of yellow and orange from FITC and that from PE. During compensation the signal arising from a specific...

...like to go to. During the conference Take notes during talks. If you have questions during the talk, jot...conference depends on what you put into it before, during, and after the meeting. Let’s break it down into...Try to avoid jargon in this initial introduction. During the conference When you meet someone new at a conference... new method that you learned about at the talk, bring it to your next lab meeting. Don’t be afraid to ...for both instances. If your poster material is nearing publication, consider uploading your work to a ...poster) so your visitors can have a DOI to go to. During the conference Arrive at the poster session early...the board and meet your poster session neighbors. During the session, if someone comes up to my poster, ...

... from Meiring et al., 2022, under CC-BY 4.0 license. A naturally occurring microtubule-severing protein...cells with physical structure, separate chromosomes during mitosis, support transport of organelles and vesicles...localized microtubule disassembly, Opto-katanin (Meiring et al., 2022), that overcomes many of those limitations...Opto-katanin induces fast, localized microtubule severing. A) Schematic of Opto-katanin constructs and activity...highly localized, rapid and reversible microtubule severing and has very low dark-state activity and toxicity...can be critical when, say, the gene is essential during development and a knockout is not feasible or when... One great way to do this is to use CRY2-CIB1 clustering. Fuse either CRY2 or CIB1 to your protein of ...

...This year, the IMP mentoring program extends its scope to the fields of Engineering, Economics, and Philology...internationalization and there is a lack of expert mentorship during undergraduate and graduate studies. Students also...Mentor Circle Programs at BWH and providing career mentoring to several students from Spain and all over the...in several countries, however programs like IMP, bring closer than ever (although not physically) the ideas...exist outside their immediate frontiers. Moreover, sharing a culture/country of origin with an international...2014-15, University of Valencia): “It is very inspiring to be in contact with mentors who are at a professional...the ability to accept feedback and adapt their mentoring styles to the mentee’s personality and professional...

..., try lowering the voltage and using a longer run time.   Samples can run into empty neighboring lanes... two most common (non-technical) issues with transferring are smaller proteins going completely through...through the membrane or larger proteins not transferring completely onto the membrane. If you’re not sure ...sure if your protein of interest is over-transferring (going completely through your membrane to the other...other side), you can check by using two membranes during your transfer instead of one. If you see signal...both membranes, you know your protein is over-transferring! If you see very little signal, but you know... sample of a large protein, you may be under-transferring.  Membrane pore size Membranes come in different...

...Kleckner. "Recombination, pairing, and synapsis of homologs during meiosis." Cold Spring Harbor perspectives...generate double-mutant offspring. Crossing mice allows for the production of offspring that have a combination...generate a desired phenotype Instead, genetic engineering should be used to separately introduce each mutation...modification will be passed along to all of the offspring in the F1 generation, your breeding scheme should... two mutations that you want combined in your offspring. Genotyping involves getting a small piece of ...is homozygous for a different mutation, the F1 offspring will be either hemizygous or null at one locus...mice that were bred in your colony, as opposed to ordering control mice from a stock center. This serves ...

...tools available for monitoring how both the brain and visceral organs interact during various behaviors.... This is all an impressive engineering feat, but you’re probably wondering “did it actually work”? The...Neuromodulation During Free Behavior. 2020 42nd Annual International Conference of the IEEE Engineering in Medicine...continuously monitor it. How to address this? Through the pairing of materials scientists (Anikeeva lab) with gut-brain...physically hooked up to invasive, cumbersome systems during experiments – neither ideal for the intrepid researcher... fiber (Park et al., 2019). This provided an engineering road map of sorts, giving an idea of the stiffness...affect their function (the gut twists and contorts during the digestion of food, while brain tissue is relatively...

... professor or hiring manager. Otherwise, use a gender-neutral greeting like “Dear Hiring Manager.” Highlight...If you are job searching, you may be wondering if you need to go to the effort of writing a cover letter...requires ddPCR skills. Your resume lists ddPCR, so the hiring manager knows that you have ddPCR experience. In...assay to answer an experimental question. Now, the hiring manager knows that you can design experiments with...briefly explaining a gap in your job history or sharing relocation plans. They can be skills-based: if ... of the department] at [name of the university].During my thesis work on [phenomenon X], I read several...joining your team as a postdoctoral researcher. Dear Hiring Manager, I am writing to apply for [title of the...

...of “bringing tomorrow’s science into today’s classrooms,” BioBuilder answers the call by offering a variety...Tamer from BioBuilder, an innovative nonprofit bringing the study of synthetic biology into the hands ... Kuldell from MIT’s Department of Biological Engineering. Built primarily from her 20 years of college... boost interest and understanding around the engineering of biology. Participating schools are provided...BioBuilder inspires students and teachers to apply engineering principles to biological systems. Today, what...BioBuilder module allows students to explore how engineering fields rely on rapid prototyping to develop and...high school teams around the world who combine engineering approaches and scientific know-how to design/...

...and scientists - Culturing a scientific network We refer to the process of mentoring within a bin as Horizontal...and more. We are partnering with ASM and are working together with the society to bring an even more valuable...next component of Binning Singletons is when we bring all the bins together at a mixer. We call this Quorum...also sought expertise from many with experience mentoring in STEM, including Addgene’s Joanne Kamens. Expanding...Chicago.  Finding our place in networking and mentoring circles We looked at a lot of approaches scientific...scientific meetings have taken to provide mentoring. For example, Goldschmidt conference, Coastal & Estuarine...historically underrepresented groups. Through such mentoring programs, scientists can build and expand their...

...28060411. Thomason, Lynn C., et al. "Recombineering: genetic engineering in bacteria using homologous recombination...leaves a “scar” in the chromosome, sometimes interfering with expression of surrounding genes. However...Developed over the past 20 years, Lambda Red recombineering, which uses enzymes domesticated from the Lambda...enzymes, require multiple transformation and plasmid curing steps, and can generate off-target mutations. ...of bacterial toxins to facilitate efficient recombineering (Khetrapal et al., 2015). They created vectors...counter-selection which enabled very efficient recombineering.     We were inspired by this work and sought...16568824. Hmelo, Laura R., et al. "Precision-engineering the Pseudomonas aeruginosa genome with two-step...

... IPI and the Springer lab (Hao et al., 2024). In this study, we, along with the Springer lab, reported...stabilizes the extended open integrin conformation. During embryonic development, extended-open integrins ...development, and the shaping of the extracellular matrix. During immune responses, this same conformation encourages...produced from hybridoma antibodies developed in the Springer lab. Some are rabbit immunoglobulin G1 (IgG1) ... to be developed and defined. According to the Springer lab, a highly specific function-blocking antibody...   Targeting integrins in the lab The Ha and Springer labs recently showed RGD-binding integrins demanded...integrins. The Ha lab — in collaboration with the Springer lab, the Jaumouillé lab, the Waterman lab, and...

...As Addgene believes many of the topics discussed during the symposium will prove incredibly important to... that the main question they wanted to focus on during the symposium was “How many graduate students/post-docs...problems in academic research and potential solutions During the first couple of sessions of the symposium it...In fact it is this very idea that postdocs are acquiring training to become professors that is provided...estimates that postdocs are paid $16.15/h before fringe benefits, less even than the $21/h [on the low ...low end] that grad students are paid before fringe benefits); what they don’t make in wages, they make up...programs as the standard option for scientists by requiring graduate students to pay for part of their tuition...

...cooking competition featuring a selected ingredient. Anyone who wants to participate brings in a dish that .... The four of us would eat together every day, sharing ideas for the company, but also talking about sports...a culture of openness and trust that comes from caring about your colleagues on a personal level, and ...special occasions. However, employees regularly bring in baked goods and we have a few unique lunchtime... from the software folks and started Hack Days. During Hack Days, teams at Addgene will forgo their regular...website. Free lunch is provided for participants during the Hack Days, and we’ve seen great success at ...university departments to establish, with the goal of fostering better communication between labs. Scientists ...

...naturally occurring Cas9 diversity via the Integral Microbial Genomes (IMG) database, Harrington et al discovered...are available from Addgene. Happy CRISPRing! References Harrington, Lucas, et al. "A thermostable Cas9...Yes Longevity in Serum?* No Yes Table 1. Comparing SpCas9 and GeoCas9. *For in vitro Cas9 RNP activity...temperatures much warmer than SpCas9 can tolerate. Harrington et al tested GeoCas9’s love of heat in two ways...can introduce genomic edits in mammalian cells. Harrington et al tested GeoCas9’s editing skills by transfecting...had similar levels of editing. See figure 4 in Harrington et al for more examples of GeoCas9’s editing ...sleeve is its increased longevity in human serum. Harrington et al suspected GeoCas9 might have higher stability...

... once a week for several months at a time during the spring and fall semesters. Our scheduling was in ...in this situation, considering stopping (temporarily or permanently) the recurring meeting or perhaps ...Enter: the journal club.  A journal club is a recurring meeting amongst scientists that aims to help stay... At my former academic department, we had two recurring journal clubs on Immunology and Microbial Pathogenesis...club? Where will the journal club take place? Preparing for the first meeting There are many important...Addgene bioinformatics journal club! What happens during journal club? There are many common ways to run...meeting using teleconferencing software. Any file sharing service should also serve to share the materials...

...in more genome engineering technologies? Browse others on Addgene's Genome Engineering Guide. Fire ...libraries expand upon the CRISPR family of plasmids by offering a lentivirus-based mechanism for sgRNA delivery...protein and cell ageing, as the spectrum shift occurs during Timer protein life due to a form of oxidation (...transcriptomes, as well as further optimization through engineering, Ed Boyden’s lab has identified a variety of ...field of optogenetics. Using a rational protein engineering approach, they designed Opto-RTKs, which activate...flavin cofactors and dimerize on exposure to light, bringing the intracellular kinase domains into contact ...activity levels and limited sensitivity under the stringently low expression conditions necessary to avoid ...

...to build a bench mentor-mentee relationship and bring your best self to the training experience.  The ... isn’t discussed often enough is the hands on mentoring you receive after you’ve found your mentor, though...lab (and possibly much more, depending on the mentoring style within the research group). The bench mentor-mentee...you don’t do proper preparation and pay attention during training, then you won’t be very proficient afterwards...how to prepare and things to keep an eye out for during training. Preparation before bench work Are there... something then you’ve done previously) you can bring those questions to your training! All of these things... the in-person training component. Documenting during training It’s hard to recall a lot of details, ...

...purchasing a gift card to use later or ordering take-out) or volunteering to foster a pet, for example. If ...is based in the lab. If you’re unsure what to do during this period, check out these tips and resources...time for exercise. Many studios and gyms are now offering online classes and participating in these is a...class twice a week at Addgene HQ, but now we’re sharing links to online yoga classes. Get outdoors. Taking...ride is a great activity for both exercise and clearing your mind. And because the outdoors is so huge...staying away from others.  Meditate. Meditating during the day is a great way to take a mental break, ...activities like team lunches, a virtual soup lunch, and sharing photos of our pets. #SocialDistancing may send...

...involves a different skill set: mentoring, setting strategic goals, and fostering team success. This shift requires...wet lab wasn’t the right path for me. I began exploring alternative career trajectories within the industry... reinvent myself and redefine the value I could bring to potential employers. Over time, I discovered ...broader vision rather than the day-to-day details. Ignoring individual differencesManaging a team isn’t “one...motivations, strengths, and learning styles. For example, during my time as a lab scientist, I worked with a diverse...frustration, micromanagement, and disengagement. By tailoring my leadership approach to each person, I was able...company by taking on responsibilities such as mentoring a junior colleague. Guiding an intern, in particular...