... "traditional" restriction enzymes in that they cleave outside of their recognition sequence, creating... BbsI), that are located on the outside of the cleavage site. As a result, these sites are eliminated ...a few advantages over other cloning methods. Exonuclease-based methods like Gibson assembly require 20...multiple DNA fragments to create TAL effector nucleases. These plasmids were designed to utilize the BsaI...

...week could expect to see their salary raised to at least a new legal minimum of $47,476 per year, under updates...universities with the FLSA in our online resource leading up to this date, as discussed in a previous Addgene...planning unionization at their institutions. One clear message that institutions have sent, with the lack...the Advocating for Science Symposium and Workshop Learn about a Variety of Science Careers Resources on...

...does it play in modern cloning, and why should you learn more about it? Read on to find out how ccdB can ...restriction enzyme-mediated disruption of ccdB, leading to positive identification of desired clones. ...most appropriate method for your own experiments. Please check out Addgene’s Choosing a Molecular Cloning...CcdB protein induces efficient ATP-dependent DNA cleavage by gyrase.” J Mol Biol. 1993 Dec 5;234(3):534-...

...Addgene blog Learn about other CRISPR methods for bacterial genome engineering   Learn about human CRISPRko...environmental isolates. For example, Peters and colleagues used Mobile-CRISPRi to knockdown individual genes...-CRISPRi. As a proof of principle, Peters and colleagues constructed a 40-member library targeting 20 ...

... suited for particular applications. Read on to learn more about each reporter or check out table 1 for...nanobody that would make a useful RANbody, consider leaving a link to its sequence in the comments below. ...29440485. Additional Resources on the Addgene Blog Learn more about using the secondary nanobody toolbox ... plasmids can be found here Watch this video to learn more about nanobodies Additional Resources Looking...

....) Fab fragments are created through enzymatic cleavage of the variable region (the antibody region that...to the antigen) from existing antibodies. Each cleaved antibody will produce two ~50kD Fab fragments, ...fragment but created through engineering instead of cleavage. They are comprised of the variable heavy and ...Whatever affinity reagents you use, one thing is clear: it’s an exciting time to be a scientist looking...

...This biosensor changes fluorescence based on the release of glutamate from neurons and this new version ...than its predecessors. Currently, Aggarwal, and colleagues Filip Tomaska and Tim Hanson, are characterizing...iGluSnFR3 in vivo and have used it to detect glutamate release from hundreds of synapses at once. “This work has...Aggarwal.In addition to the biosensors, Aggarwal and colleagues are working on developing new surface display...

...unfortunate experience of witnessing…nothing. For some unclear reason, cells are not getting transduced. Here,...reducing the culture volume into which the virus is released, a more concentrated virus solution can be obtained...PMC330758. Additional Resources on the Addgene Blog Learn All about Adenovirus Associated Virus New to Virus...Virus? Check out Our Primer for Beginners! Learn All about Lentiviral Vectors Additional Resources on ...

...target DNA without cleavage until it's activated by light. Once activated, Cas9 cleavage can induce DSBs ...probes and editors. A probe array may consist of at least two sensor modules targeting a specific chromatin...complex binds to the degron sequence ultimately leading to the degradation of the AID-tagged protein. To...

...spectrum. The experimental apparatus should be cleaned thoroughly between animals as olfactory stimuli...animal (ex: urine) can be distracting. If it is not cleaned, animals tested towards the end of an experiment...coordination to more complex situations such as learning and memory or emotional states, for example anxiety...Guide to Viral Vectors Resources on Addgene.org Learn about Addgene's viral service Find viral protocols...

...adenovirus, and adeno-associated virus (AAV). The least risky of these four viruses is AAV – it is not known...propensity to integrate into proto-oncogenes, which can lead to cancer. These viruses need to be handled in a...thoroughly decontaminated after use, typically with bleach (plastic) or ethanol (other surfaces). For AAVs...and isopropyl alcohol, 0.5% peracetic acid, 10% bleach, or 1% iodine are useful. Some organizations or...

...Arabidopsis in Education Learning Synthetic Biology with BioBuilder 3D Models for learning about CRISPR Resources... subsequently destroyed by Argonaute-catalyzed nuclease activity or their translation disrupted through...protocols and want to share pictures, feedback etc. please write to [email protected] and if you make...

...Addgene Blog Learn about Synthetic Biology with BioBuilder Use Addgene Videos to Learn about Basic Lab...genes in other species, but these studies generally leave questions about gene function wide open. What if...Check Out Our Molecular Biology Reference Pages Learn Simple Lab Techniques in Our Protocol Videos ...

...the promoter region of gamma hemoglobin (HGB1/2) leads to continuous expression of fetal hemoglobin even...efficiency, respectively) and outperformed ABE7.10 by at least 1.4 fold. The ability of ABE8.20-m to edit primary...editing of A•T to G•C in genomic DNA without DNA cleavage. Nature 551:464–471 . https://doi.org/10.1038/...Addgene blog Find more blog posts about base editing Learn about another evolved base editor: ABE8e Read about...

...serotype, as some rAAV serotypes are abundantly released into the culture media, while others remain cell-associated...and non-genome-containing (empty) capsids, thus leading to an overestimate of transducible units in the...and dehydration (Howard & Harvey, 2017). You can learn more about common AAV applications by reading our...Wilson, J. M. (2010). Efficient serotype-dependent release of functional vector into the culture medium during...

...desired recombination event will remove NeoR only and leave exon 2 floxed, as seen in line 4 of Figure 4. Since...tools for creating knockout animals or cell lines, please consider sharing them by depositing with Addgene... with CRISPR through Non-Homologous End Joining Learn How the FLEx Switch Can Be Used to Turn Genes On...Mammalian Expression Vectors Find Cre-Lox Plasmids Learn More about CRISPR  ...

...remember what to say and provide your audience with a clear picture of your work. Practice, practice, practice...with people inside and outside your field. Your colleagues, lab mates, and PI can provide feedback on your...Anticipate questions: When practicing with your colleagues, ask them to brainstorm potential questions you... of a member of the Addgene depositor community Learn about careers at Addgene ...

...communication. But first, take a deep breath; at least you know you have an answer to that previous nosy... complicated process even more difficult. I’ve learned that some of the technologies that helped us navigate...application process and usually occurs in the few months leading up to applications opening. With virtual options...be a great opportunity to introduce yourself and learn more about your schools of interest. Benefits of...

...four times in a 95% ethanol and dry ice bath to release the virus particles from the cells. Cell lysates...batch-to-batch variability. Check out this post to learn more about different ways to titer AAV. Do you ...you have any AAV production tips or tricks? Please share them in the comments below! Special thanks ...first time CRISPR user Resources on Addgene.org Learn how Addgene produces AAV and lentivirus Check out...

... blog Learn the basics of Golden Gate assembly Check out NEB's work on ligase efficiency Learn about ...assembly, ligation of mismatched pairs of overhangs can lead to incorrect assemblies, so care must be taken to...is a molecular biologist and a Development Group Leader overseeing a team bringing new products to market...