...toolkit New Cas protein identified Jennifer Doudna’s lab recently identified Cas14, a family of small...or DNA. Now they have developed this further to provide multiplexed detection, higher sensitivity, and ...of DNA Elements (ENCODE), a consortium aimed to identify functional elements in the human genome, these...sheet Resources on Addgene.org Read our CRISPR guide Find CRISPR plasmids by function Find recently deposited...

...scientists outside of the US now use the repository – about 50% of plasmids are shipped to labs outside of the...plasmid shipping look like at Addgene? Here's an inside peak into the daily workings in Addgene's lab. ...world... Where did 500,000 plasmids go? Far and wide – that's where! Check out the map below which depicts... improve the repository. We love being able to provide a useful (and growing) resource for our favorite...

...Plasmids 101 videos and needed a break so they're hanging out with Aavery in this Zoom video background....background. Want the purple bacteria video background with mascots Blugene and Aavery? Download the 720p version...our virus backgrounds. Addgene’s viral service provides ready-to-use AAV and lentiviral preps! Download...DNA background. Or get the same background as a video background! (720 px or 1080 px).         Do you...

...library screening, read our recent blog post on Genome-wide Screening Using CRISPR/Cas9. If you need help cloning...v2, you can take a look at the protocol kindly provided by the Zhang lab. Even though lentiCRISPR v2 has...of matrix metalloproteinase-9 secretion, and identifying lysine methyltransferase substrates. We’re incredibly...Shalem, and Feng Zhang. "Improved vectors and genome-wide libraries for CRISPR screening." Nature methods ...Resources on the Addgene Website Visit our Lentiviral Guide Pages Find Lentiviral Plasmids Find CRISPR Pooled...

...when enhancers were active or silent, they could identify the target genes of previously uncharacterized...has made targeting much easier! With the help of guide RNAs, fusions between Cas proteins and the epigenetic...this fusion protein allows for LSD1 targeting to a wider set of genomic targets. These two vectors are part...CRISPR-based DNA demethylation by Casilio-ME-mediated RNA-guided coupling of methylcytosine oxidation and DNA repair... the Addgene Blog Cas9 Activators: A Practical Guide CRISPR Between the Genes: How to Experiment with... Resources on Addgene.org Check out Our CRISPR Guide Pages Find Validated gRNAs to Target Your Genomic...

...Watakabe et al., 2015) (Figure 1). You’ll need to consider if such properties could help, or harm, your experimental...tools, where modification of cellular activity outside of your experiment site or circuit can occur. Promoters... promoters  This type of promoter allows you to decide when to turn on, or turn off, gene expression by...promoters to begin testing, you’ll next need to decide how much virus to put into your system, which is...injection volume.  There are a few factors to consider. First, the titer required is inversely related... posts in our Viral Vectors 101 series can help guide you with these, so keep reading! Good luck, and...

...there are up to thirteen natural serotypes of AAV identified so far, numbered AAV1, AAV2, and so on. In addition... the second number to the capsid. Most vectors provided by Addgene are pseudotypes using the genomic structure...AAVrg has not been thoroughly studied in contexts outside the nervous system to confirm its cellular receptors...2008). Adeno-Associated Virus (AAV) Serotype 9 Provides Global Cardiac Gene Transfer Superior to AAV1,...., Robbins, P. D., Li, J., & Xiao, X. (2006). Widespread and Stable Pancreatic Gene Transfer by Adeno-...Capsids Resources on Addgene.org AAV Data Hub AAV Guide AAV Packaged On Request Browse all in-stock AAV ...

... coming out of MITs Human Dynamics Laboratory identified five characteristics of very successful teams...this series). Members carry on back-channel or side conversations within the team. We are not talking...contribute. Members periodically break, go exploring outside the team, and bring information back. No team ...colleague: MBWA "Manage By Walking Around" – don't hide in your office. Stop by instead of emailing.    ...volunteer constructive feedback and suggestions. Consider offering rewards (gift card, afternoon off, etc...

...mostly because of this article: “A systematic identification and analysis of scientists on Twitter.” This...Scientific Disciplines on Twitter. Twitter users were identified as scientists if they had a scientific title ...scientists within their fields than scientists outside their fields Figure 6: Follower Network of Scientists...follow scientists within their field rather than outside their field. Lower: Follower networks of subgroups... Y., & Sugimoto, C. R. (2017). A systematic identification and analysis of scientists on Twitter. Plos...

... a very different schedule. Fortunately, I was provided with a detailed training schedule that helped ...it relates to other teams at the company. This provides new (and old) employees with information on how...possible to work the remainder of the three hours (outside of the core hours) at a time that works for you...transition into a traditional work environment was aided by the efforts being put in by Addgene to ensure...blog.      More resources on the Addgene blog A Guide to Getting Started in Undergrad Research Tips for...

...requesting and ordering. This split allows us to provide researchers with increased choice and flexibility... Plasmids available for Packaged on Request consideration will have a banner on their material page. Interested...testing, and viral genomic sequencing to confirm the identity of the packaged DNA and serotype. Please note ... in-stock preps. If a request has technical considerations that may affect expected titers, a customer...1–2 days (USA); 3–4 days (international; varies widely by country and/or distributor.)  You should plan...

...certainly hope not! One of our core values here is to provide unmatched customer support, as this is vital to...exact problem, and it’s called “self-service.” The idea here is to build features that allow users to help...general question or concern about the answers we provide, within the widget you also have the option to ... we’ll be continually improving the support we provide based on your valuable feedback. This is Addgene...first. Resources on the Addgene Blog Read other Inside Addgene Posts Learn about New Features on Our Plasmid...

...joined by Addgene Scientist Matt Ferenc (one of our resident CRISPR experts) at the 2014 Johnson & Johnson’...up to the discovery of the mechanism of dual-RNA-guided DNA cleavage in adaptive bacterial immunity and...Dounda and Charpentier met at a 2011 conference and decided to collaborate on what was a “curiosity” driven...thanked Martin Jinek and Krzysztof Chylinski (who coincidentally speak the same Polish sub-dialect making their... Technology? Learn More: Addgene's CRISPR/Cas9 Guide History of CRISPR Cas - A tale of survival and evolution...

...biosensors When choosing a biosensor, you first need to decide the best way to track or measure the process you... of small molecules like arabinose or hydrogen peroxide or ions like calcium and zinc. Some biosensors...compatible with your lab and experimental setup? Consider what kind of instrumentation you have available...components of individual cells Separation/identification of fluorescently-labeled components Fluorescence-activated...Optogenetics Plasmids Visit the Fluorescent Protein Guide Pages ...

... mapping where a protein binds genome-wide, and has been widely applied in many model organisms and cell... Enter CUT&RUN, an enzyme-based protocol for identifying DNA binding patterns that overcomes some of the... degree of signal to noise is desired. It also provides an excellent additional measure of DNA binding...technique will quickly become a standard method alongside ChIP. Many thanks to our guest blogger, Matthew...

...clinical data about a disease or therapeutic Side-by-side comparison of the same experiment from different...makes your insights discoverable. Watch this quick video to learn how ReFigure works and continue reading...Apps to Save Time and Track Your Experiments Use video to share your science at We Share Science Learn ...with Addgene Find Molecular Biology Protocols and Videos Find Other Molecular Biology Resources ...

...post-grad plans, but in the end, writing won out. “I considered teaching at a liberal arts school but I’ve always...Iowa, but kept up a freelance writing hustle on the side. “I actually stumbled upon, about 6 months in, this... was for Cactus Communications, a company that provides scientific editing services. “I was working during...At the end of her postdoc, Caitlin had a clearer idea of her career goals: “My plan was to go into editing...landing her job. All of the hard work she’d put in outside the lab had paid off. “The combination of the editing...

...Visit our viral vectors guide Find our chemogenetics and optogenetics science guides ...Chemogenetic approaches excel at broad tissue or system-wide targeting, as the ligand can reach target cells ...optogenetics. If broad control of whole circuits or tissue-wide systems is necessary, use chemogenetics. Note that...approaches suffer from unique sets of off-target side-effects that should be carefully controlled for ...

...for your CRISPR genome engineering experiments, consider that wildtype Cas9 with optimized chimeric gRNA... drive expression, is it necessary to add a G nucleotide to the start of my target sequence? A2: The human...screened? A4: Before starting your experiment, consider co-transfecting with GFP. This allows you to sort... FAQs answered by Le Cong Read Addgene's CRISPR guide for background information on CRISPR/Cas9 systems...plasmids Links to different software to help you identify your gRNA target sequences Additionally, I would...

...demonstrate that 85% of leaving postdocs pursue a career outside the traditional academic silo to tenured professor....  Here are some questions and perspectives to consider. What do academic and non-academic careers in ... a greater sense of autonomy - there are fewer outside influences on your topics of research or course...move around every 3-6 years—a short and exciting ride Complicated matrix structures are common Small... a scientist you enjoy the most. Having a better idea of where you want to go will help you focus on the...

...useful, chemical fluorophores can still be used in a wide variety of experiments. Read more to find chemical...potential applications of R-bodies and to find a video of them in action. AAV: A Versatile Viral Tool for...interest to a variety of cell types in-vivo. These are widely used by the optogenetics and chemogenetics communities...pathways using light and chemicals respectively. Didem Goz Aytruk from Connie Cepko’s Lab at Harvard Medical...AAVs function and indicates important things to consider if you’re getting started with them. CRISPR Pairing...

...resources and experience to their advantage should they decide to produce virus on their own.Harvard researcher... What viral preps are available now? There are a wide variety of tools already available from Addgene ...be continually expanding our collection. We’ll provide a brief rundown of what lentiviral and AAV preps...it, modulate its expression, or perform a genome-wide screen, we’ve got viruses to help you out. Of course...the Addgene website. blog.addgene.org The blog provides tips for working with viruses including advice...

...developed and tested by ABRC staff. These kits provide in-depth, ready-to-teach lab protocols and supporting...supporting materials such as instructional videos and datasheets. The remaining kits were donated to ABRC...cycle, and is relatively easy to grow, it is an ideal organism for teachers looking to expand their science...outlined in ABRC’s kits vary by module. As a general guideline, experiments that include growing plants require...available via e-mail and phone to answer questions and provide support. Growing capacity - ABRC outreach today...

...service that we, as scientists, provide for journal editors to help decide whether work is suitable for ...of the 18th century. Yet the peer-review was not widely adopted by the scientific community until the mid...peer-reviewing journal submissions since 2015 for a wide variety of journals. Since then, I developed a routine...submitted paper in addition to the journal-specific guidelines.  Peer-review step 1: The first read At the first...

...through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ...a new CRISPR screening method that makes use of guide RNAs with internal barcodes (iBARs) in their loop...experimental replicates for each of the gRNAs. The idea was to assign four different barcodes to each of...covers every annotated human gene, it enables genome-wide screening with a high MOI, which is beneficial for... and can serve as a drug-screening platform to identify new antifungals that target calcineurin. Current...

..., Addgene will have unique Research Resource Identifiers (RRIDs) for each antibody, making it easy for...topics are discussed on Addgene’s blog and Antibody Guide, which include information about immunofluorescence...suggestions about how to make the resource better or ideas for new materials to add to the collection, please...find useful antibodies for your research. Please consider sharing your experiences with antibodies from ...Introduction to Immunofluorescence Addgene’s Antibody Guide The project is supported by a NIH BRAIN Initiative...

...transformation fall into two main categories: antibiotic/herbicide resistance and fluorescent proteins. Selection...undifferentiated plant cell culture), which is great for a wide variety of applications. But some techniques require...CRISPR-Cas9 gene editing, transformation may be identified by resistance to an antibiotic. Successful editing...therefore, are useful for research, and they are identified by a phenotype caused by the segregating mutation...these mutant plants. Using RUBY to simplify the identification of these mutants will speed up my work and ...

...important to use a high fidelity polymerase to minimize mutations. The fidelity of the polymerase becomes...as Addgene’s Sequence Analyzer, to allow you to identify which restriction sites are present in a given...compatible with a standard PCR reaction (see PCR Video). Therefore, our Forward Primer will use the sequence... chance for mis-priming, so you can use a pretty wide range of annealing temperatures, but you may need...

...this stage, only cells from the immune system show wide scale mobility with movement from the bone marrow...you can detect signal in thick specimens. See the video below for an example of two-photon intravital microscopy... in diverse tissue environments. These studies provide quantitative measurements of cellular motility,...and skin. Two-photon intravital microscopy. The video above depicts the infiltration of neutrophils expressing...technical specificities have to be taken into consideration especially in the use of fluorescent proteins... Addgene.org Check out Our Fluorescent Protein Guide Pages Find Plasmids for Subcellular Localization...

...scarless DNA constructs using homologous regions to guide the joining reaction. BioBrick Assembly will leave...fragments, and generate constructs for producing single-guide RNAs – but it also allows you to skip purification...NEBuilder HiFi so accurate (apart from the high-fidelity polymerase) is the ability to remove, reset and...freely available online. Its simple interface can guide you through both experimental and primer design ...single-stranded synthetic oligo to generate a single-guide RNA (sgRNA). Using only one single-stranded oligo...

...Nicholas T., et al. “Genome-wide analysis in vivo of translation with nucleotide resolution using ribosome...sequenced and aligned to the genome to determine genome-wide translational frequency. Although highly useful,...reporter mRNAs. Using translation inhibitors cycloheximide and puromycin, Halstead et al. verified that... translated, the SunTag sites on the nascent polypeptide recruit scFV-GFP, generating green puncta that...

...smaller fragments that are then barcoded for identification. This mixture of barcoded fragments is known...for each sample remain highly normalized across a wide range of input DNA concentrations. This process ...of 96 plasmids on a plate) will typically have a wide range of input quantities. This variation is problematic...are produced from large numbers of samples with a wide range of input concentrations (via proprietary chemistry...alternative to Sanger sequencing because the sample identity information is always displayed at the individual...

...working in academia or industry. At BosLab, we identify ourselves as part of a network of independent ...square, but we saw the potential in the space and decided to reactivate it. We started out by holding a few...from hacking probiotics to filming educational lab videos, while some are looking to develop new skill sets... mind. Thus, to encourage more involvement and provide all our members the opportunity to do molecular...project. This project was inspired by previous city-wide genomic studies and our study will involve getting...

...product because the plasmids in the library are identical except for the insert; there’s no need to sequence...results represent a successful amplification? To provide a point of reference, Addgene commonly prepares...results If your sequencing is successful, your NGS provider should return results in the form of many, many... want! At the end of the day, it’s up to you to decide your threshold of what a good amplification looks...post has helped clarify some of the important considerations when using, amplifying, and verifying pooled...

... the widely used 293T. During vector production, assembled AAV vector particles accumulate inside the ...either associated with the outside surface of the exosome or are carried inside the vesicle, which may also...Addgene's viral service Find viral protocols and videos Find AAVs for your research ...

...physical location didn’t matter. I didn’t have to consider that aspect and I could plan more with regard ...depending on the conference, the audience can help decide which questions get answered. “You can upvote a...presenting is eye contact. “It’s important to keep your video on and look into the camera to make it seem more...on Twitter where each poster presenter had four “slides” to use for their poster, eliminating the need ...to make space for it,” says Bachle. “On the flip side, you can attend conferences for just a little bit...

...situation. But a few days later we had an idea. And this idea rapidly grew and became Crowdfight COVID-...modern science in our view. And it was this very idea that research is done better with others than by...has been essential to push certain projects. The idea behind the platform was simple: any person working...remote collaborations Read our Transferable Skills Guide article about external collaborations Resources...

...efficiencies and expression levels in the cells. Ideally, the chosen reporter is unique to the cell, is ... generations. Stably transfected cells may be considered an entirely new cell line from that of the original..., NIH3T3, CHO, 293HEK, Jurkat T cells Blocks polypeptide synthesis at 80S; inhibits chain elongation 100...hygB HeLa, NIH3T3, CHO, Jurkat T cells Blocks polypeptide synthesis at 80S; inhibits chain elongation. ... 10-14 days depending on how quickly your cells divide. Gentamicin is often used as a supplement in mammalian...

...suggests a change to a slide, think about whether the same principle applies to other slides in your deck.  If... craft sentences for abstracts and presentation slides, learn from audiences as they do science outreach...of a new science communication initiative that provides yet more scientists with opportunities to practice...Resources on Addgene.org Check out how Addgene uses videos for science communication ...

...in the lab. This is commonly done using calcium chloride which permeabilizes the cell membrane so the bacteria...inject their genetic material into the cell. Once inside, phages can follow one of two different life cycles...both of these life cycles bacterial DNA can be accidentally packaged into the newly created phages. Transfer...referred to as transduction. Transferred DNA once inside the infected bacterium can either exist as transient...Addgene’s protocol page, which has both protocols and videos for techniques in basic molecular biology, plasmid...

...protein. Animals are first injected with a small peptide that represents a short, unique amino acid sequence...isolate individual hybridomas and ELISA is used to identify the hybridomas producing the most specific antibodies...antibody, meaning that the antibodies are largely identical between lots.  Disadvantages of monoclonal antibodies...antibody-encoding genes over time as the hybridoma cells divide. These changes can alter the antibody that is actually...produced by the hybridoma. Any genetic drive can be identified by sequencing the heavy and light chains. To ...

...Visit Our AAV Guide Page Find Ready-to-use Virus for Your Research Visit Our Lentivirus Guide Page ...understand brain function So far scientists have identified 180 brain regions. Both this geography and the...Using retrograde AAV AAVrg can be used to deliver a wide array of genetic tools for neural interrogation ...where the cell bodies of the projection neurons reside.   Impact for scientists While the mechanisms ...

...of dNTP in the reaction mix, the researcher can decide exactly how far the T4 exonuclease proceeds: at...at the first occurrence of the chosen nucleotide, T4 will switch to polymerizing and then stall due to...the “damage” and producing a complete plasmid. Besides being compatible with any vector, the SLIC protocol...multi-fragment DNA assembly, and its low cost makes it ideal for researchers doing large amounts of cloning. ... 10.1007/978-1-61779-564-0_5. PubMed. JBEI-J5’s guide to SLIC Resources at Addgene Ligation-Independent...

...particular marker, you can use the reference plots provided by the antibody companies or check the literature...sample. The isotype antibody will be virtually identical to the antibody for your marker of interest. It...for that particular marker. This enables you to identify any non-specific binding to know that your positive...unsure what amount of antibody to use, you should consider using an isotype control. Pro tip! Many companies...Cytometry Resources on Addgene.org Addgene's Antibody Guide Browse Addgene's Antibody Collection        ...

...more OptoNBs would allow for more uses both inside and outside cells. References Gil, Agnieszka A., et...exciting because using light to control signaling provides greater spatiotemporal resolution than pharmacological...domain of the Sons of Sevenless (SOS), a guanine nucleotide exchange factor that activates Ras. They expressed...more about optogenetics in Addgene's optogenetics guide ...

.... To make a point mutation, you design two oligonucleotide primers with the desired mutation, then use...3-4 phage proteins are needed and these can be provided via a separate plasmid or be inserted into the...insertions and deletions. The figure above shows guidelines for primer design; a desired insertion should...REPLACR-mutagenesis successfully incorporated a 60-nucleotide insertion; beyond this insertion size, primer...introduce mutations. Using commercially available high-fidelity polymerases such as KOD and Phusion, which are...

... in the human genome. The pair has focused considerable attention on transcription factors, starting...have good antibodies. The results look almost identical. Addgene: What advantages does your system have... also easy to run things in parallel. Addgene: Besides being fast and efficient, what other challenges... Myers: Only a small percentage of the proteins inside a cell have a commercial antibody against them.... to us to get them. Even though we’re happy to provide them, that never works on a large scale. The world...

...to $47,476 per year. This post discusses how a nationwide injunction against the FLSA is affecting universities...injunction against the FLSA updates was granted nationwide. Again, we at Future of Research believe that...higher than the level being raised to now - across a wide consensus of academic stakeholders. However, while...collecting data on the effects of the injunction nationwide and, at exactly one month following the injunction...plans, but have since reversed their position and decided to allow individual departments to go ahead with...

...time-consuming aspects of traditional cloning is the identification of clones that actually contain your insert...enzyme-mediated disruption of ccdB, leading to positive identification of desired clones.   ...Becomes a powerful... insert of interest. Hence correct clones are identified much more efficiently, as those that do not contain...ultimately perform the same function, there is some evidence that ccdB Survival™ can be more difficult to transform...E. coli do not contain the F plasmid (and are considered F-); however, there are a few popular lab strains...

...even come [to grad school]!” He asked if she had considered other careers, which she hadn’t.  He then said... didn’t know if she liked science writing. She decided to start a blog to test it out and discovered she...communications planning.” To beef up her resume, she decided to complete a certificate in Strategic Communications...these difficult pieces: “It’s probably the best.” Besides working through the tough assignments, the skills...important for her work include: “Listening. Evaluating evidence. Critical thinking. Being able to learn things...