We narrowed to 1,082 results for: TOR

...different Cas12a inhibitors. Some of these anti-Cas12a proteins have broad inhibitory effects on Cas12a...Caixia Gao’s lab has developed a new plant base editor from an evolved tRNA adenosine deaminase that is... is fused to a nicking Cas9. This base editor can be used for A•T to G•C edits in protoplasts or plants... Biology Find plasmids containing these base editors Spatially and temporally regulated AAV-based ...single step gRNA multiplexing system that generates vectors containing high numbers of gRNAs in one overnight...overnight reaction. STAgR is highly customizable as vector backbones, and gRNA structure, numbers, and promoters...Mobile-CRISPRi plasmids Identification of Cas12a inhibitors Using a bioinformatics and a screening approach...

...zinc finger nucleases to transcription activator-like effector nucleases (TALENs). Our most popular kit... Our first popular "kit," Andrew Fire's set of vectors for C. elegans research was our most requested ...expression, such as pLKO.1 puro and pLKO.1 – TRC cloning vector. Lentiviral Plasmids -  If you're familiar with...requests grow for lentiviral packaging and expression vectors, including pMD2.G and psPAX2. Actually, these two...now have 19 different ubiquitin plasmids in the repository. 100,000 Plasmids Shipped! - By 2010, Addgene...be popular years after they were added to the repository, including beta-catenin reporter M50 Super 8x... GCaMP6 series of genetically encoded calcium indicators were the hot technology of the year in our collection...

...DNA-binding domains based on transcription-activator-like effectors (TALEs) as anchors for genome engineering... “Transcriptional activators are easy to expand or combine with different effectors. I’d encourage people...The results are light-inducible transcriptional effectors (LITEs) designed to bind specific genes and turn...devised the plasmids now available in Addgene’s repository for use in neurons, both in culture and in living...Zhang showed they could package LITEs in viral vectors and genetically target them to specific cell populations...expression levels reliably increased in vitro by a factor of 10 to 20. A flexible and modular optogenetics...

...NHEJ). Designing an RNA editor The Zhang lab envisioned a two-component RNA editor: a Cas13 enzyme fused...after that, the lab published a paper on an RNA editor that allows C to U edits (RESCUE). We’ll walk through...translational machinery treats like guanine. This RNA editor would permit point mutations in RNA which could...nonfunctional. In their quest to build a robust RNA editor, the lab started with the Cas13 scaffold, testing...PspCas13b. Find the plasmids for the RESCUE RNA editors here! Testing and improving RNA editing from the...is too large to fit into adeno-associated viral vectors, they tested truncated ADARDDs to see if they could...edit multiple RNAs. It’s clear that CRISPR RNA editors will open new doors for scientific research - how...

...molecules can alter the overall 260 nm reading, distorting the final concentration calculation and potentially...(Figure 1B). The A260/230 ratio can be a good indicator of protein contamination, but a number of other...contaminants alter this ratio as well (Koetsier and Cantor, 2019). The 280 nm peak is more specific to proteins...researchers traditionally use the A260/280 ratio as an indicator of protein contamination. However, this ratio ...salts, and detergents (Figure 1C-D; Koetsier and Cantor, 2019). Guanidine salts and detergents have absorbance...at concentrations as low as 10 µM (Koetsier and Cantor, 2019). As such, a sample with a good A260/280 ...the presence of RNA contamination (Koetsier and Cantor, 2019). To confirm RNA in your DNA sample, you ...

...papers are analogous to mammalian viral vectors in that the vectors are replication-deficient. Thus, a very...like for organisms like C. difficile. In 2014, Citorik et al. and Bikard et al. developed sequence-specific...antimicrobial, plasmid-based, CRISPR systems (Find the Citorik Plasmids here). In both cases, they used 1-2 CRISPR...sequences found only in certain bacteria (Figure 1). Citorik et al. targeted beta-lactam and quinolone resistance... for bacterial CRISPR targeting. Phage-derived vectors carry CRISPR RNAs (crRNAs) targeting either genomic...suggest that targeting multiple resistance/virulence factors may further reduce this likelihood. The key problem...hardest-to-kill bacteria. In using phage-derived vectors to carry CRISPR, we’re turning the tables on the...

...Expression Vectors Find Bacterial Two Hybrid Plasmids Read our Blog Post About Yeast Vectors Read our Blog...discovered the modular format of some transcriptional activators: that the DNA binding domain (DBD) and transcriptional...of whether it interacts with a transcriptional activator (1a) or a repressor (1b). Scientists took advantage... original Y2H systems utilized the yeast Gal4 activator, bacterial LexA DBD and the lacZ reporter gene...detectable reporter signals depend on a number of factors, including protein expression and correct folding... is to use a number of different bait and prey vectors. This has been shown to be as effective as using...transcription-activating function of a eukaryotic regulatory protein. Science. 1986; 231:699-704. PubMed PMID...

...This post was contributed by Doug Richardson, Director of the Harvard Center for Biological Imaging and...block out of focus fluorescence from reaching a detector or restricting fluorescence to specific excitation...light passes through a pinhole before reaching the detector. The pinhole is specifically placed to only allow...allow light from the focal plane to reach the detector and all light from above or below is excluded. ...the optics of a microscope before it reaches the detector. This means that two or more objects lying within...collected and projected back to a fast, sensitive detector such as a scientific CMOS camera. If plasma membrane...higher photon flux via the more efficient camera detector and lack of a light-reducing pinhole. By collecting...

....com. Mentors You may have an adviser, but do you have a mentor? There is a difference. A mentor is someone...that many graduate and post-doctoral programs lack any formal or mandatory professional development training... you may need to look a bit harder to find your mentor. This may be a connection you’ve made at a conference... Alternatively, it might even be a colleague. A mentor does not necessarily need to be someone a lot older...planning, has written a great blog on choosing good mentors for scientists. Continuing professional development...popular job site webpages. Importantly, find a good mentor that can share relevant experience and resources...

...place. As a curator of the thousands of amazing plasmid technologies contributed by our depositors, Addgene...plasmid" section Anytime you use a plasmid from the repository, we request that you cite it using both its Addgene...increases the visibility of both you and the original depositor. Our citations feature makes it so that, when ...reliability of our search results depends upon you, our depositors, giving us as much information as possible during...you to browse through the many plasmids in the repository, our scientists have curated a number of special...attention to the great variety of plasmids in the repository. And last, but certainly not least, we often ...Special Collections Pages Suggest Plasmids for the Repository Find Plasmids for Your Research  ...

...Cytosine base editors with minimized off-target DNA editing The original cytosine base editor, BE3, induces...deamination. They generated these editors using the BE4max architecture. The new editors reported in this study ...the paper in Nature Biotechnology Find the base editor plasmids CRISPRi in Caulobacter crescentus ...ctrA, a gene encoding an essential cell cycle regulator. The lab expressed both dCas9 and the ctrA sgRNA...lab developed new fluorescently labeled piggyBac vectors to deliver uniform and sustained expression of ...editing by TET1 by co-delivering TET1 and protein factors for linking methylcytosine oxidation to DNA repair...

... chimeric antigen receptors (CARs). CARs are a chimera of scFvs and T cell receptors and are expressed...portion of the receptors recognizes an antigen of interest while the intracellular T cell receptor portion of...domains of the selected scFvs are inserted into a vector encoding the antibody constant domain scaffold ..., and enzymes for cancer, autoimmune, and/or inflammatory therapeutic applications. Antibody fragments...cancer-related antigens, binding of the scFv part of the receptor can activate T cells to kill cancer cells. scFv...small size also allows for their delivery by viral vectors like AAV, which could be useful for delivering ...doi.org/10.1038/nbt.1791  Deal CE, Balazs AB (2015) Vectored antibody gene delivery for the prevention or treatment...

...papers or are brainstorming new technological innovations. Let’s take a look at three historical techniques...that save both time and money. Yet, knowledge of historical techniques comes in handy whether you are perusing...real-time PCR Southern blotting, named for its inventor Edward M. Southern (1), can be used to detect ...labeling it. The label can then be visualized by autoradiography and X-ray film for radioactively labeled probes...DNA technologies that are now commonplace in laboratories. As scientific technologies rapidly progress...throughout the world. Let us know your favorite "Historical Lab Technique" in the comments section below!...Coulson. "DNA sequencing with chain-terminating inhibitors." Proceedings of the National Academy of Sciences...

...in the specific repositories (Addgene, WormBase, Arabidopsis.info). These repositories will ensure that...post was contributed by Magdalena Julkowska, a postdoctoral researcher at KAUST, Saudi Arabia. From the...service that we, as scientists, provide for journal editors to help decide whether work is suitable for publication... submission to the subsequent journal.  As a postdoctoral researcher studying plant development and abiotic...the data can be deposited in the public on-line repository (like GitHub, Zenodo, Figshare, Triad) while ... and Records: The Royal Society Journal of the History of Science 69.3 (2015): 337-352. PubMed PMID: 26495581...protocols Find eBooks on plasmids, CRISPR, viral vectors, and more ...

...fully-equipped laboratory for citizen scientists. In addition to maintaining a laboratory to support both... of independent Do-it-Yourself (DIY) biology laboratories working towards enabling everyone to participate... set on top of filing cabinets, a rickety old incubator, a few pipettes, and some home-made equipment....Anyone can become a member of BosLab and previous laboratory experience is not required. Our members are skilled...have non-traditional scientists working in our laboratory! Don’t worry though, we make sure that everyone... became our bacterial transformation guru and instructor for our intro to biohacking courses. Our members...a co-organizer at BosLab, a community biotech laboratory in Somerville MA. Follow her on twitter at @ZebraElement...

...therefore we are not a party (signatory) to the MTAs for plasmids in our repository. Instead, we facilitate ...and Recipient (in Addgene parlance, the depositor and requestor) organizations. Over the last twenty years...with the number of requests and deposits in our repository. From 2004 to 2023, we facilitated 784,795 MTAs...MTAs per year.  The MTA Process at Addgene As a repository, Addgene doesn’t actually own the majority of...your organization agrees to let Addgene receive, store, and distribute your material under the specified...move forward with shipping the material to the requestor.  Why the MTA?  The MTA process can be a bit confusing...

...licensed distributor) allow you to quickly transfer a DNA sequence from the transposon vector to one of...encourage you to deposit your piggyBac™ transposon vectors with us to help us expand this useful resource....such as with signaling molecules or transcription factors, or when the gene is constitutively-active or a...edit of interest. With the larger piggyBac™ donor vector, it is possible to make the ITR and selection cassette...re-cutting. Transposase removal of the cassette later restores the target sequence. This allows researchers to...Transposase Read up on Genome Engineering with FLEx Vectors  Learn how Homology Directed Repair Is Used for...Genome Editing Resources on Addgene.org Find Vectors with the piggyBac™ Transposon Catch up on Everything...

...reporters require a chemical substrate (and sometimes cofactors) to be provided and are generally dimmer than ...Kinetics of photon generation Slower Faster Cofactors/substrates Required Not required Signal strength...engineered Peak emission (nm) Size (kDa) Substrate Cofactors Secreted North American firefly (Photinus pyralis...AkaLumine ATP No Luciferase reporter assays for monitoring gene regulation Since firefly luciferase was ...expression analysis to investigate the effect of regulatory elements — such as promoters, enhancers, or untranslated...untranslated regions — on gene expression. The regulatory element is cloned upstream of the luciferase ...energy from an excited donor fluorophore to an acceptor fluorophore, which then emits it as light. Bioluminescence...

... DNA sequence called the Fertility factor (F-factor). The F-factor is found on an episome, a piece of ... information on viral vectors, including transduction download our Viral Vectors 101 eBook. Bacterial ...fragmented DNA released into the environment. Various factors promote natural transformation in different bacteria...bacterial or eukaryotic cells via a virus or viral vector. One example are bacteriophages that attach to ...scientists commonly use phagemids, a DNA cloning vector that contains both bacteriophage and plasmid properties...

...entire mission is to enable that process. Our repository of plasmids, AAVs, antibodies, and data allows...technical skills, and experiences from thousands of laboratories that makes up our Help Center, blog, website... the blog originally started as a way to easily store and share in-depth answers to frequently asked questions...on a case-by-case basis, as many of our blog contributors primarily write in the areas in which they have...graphic designer upon request that the science communicator "just sketch a western blot out quickly." Final...and cited figures from papers or image sharing repositories. Then we format, do a final review to catch ...informative resources that answer questions our depositors and customers are asking. So how can you get ...

...contributed by Maaike Pols, PhD, a Developmental Editor at F1000Research.  With many changes taking place...their funding should also be published. All these factors will influence your choice of journals to submit...communities. The ARRIVE guidelines for the reporting of laboratory animals during in vivo research and the CARE ... are two sets of guidelines frequently used by editors. Providing all this information makes your manuscript...who the reviewers are). If the reviewers and the editor decide that your manuscript needs to be revised...the conclusions of your paper. This will help the editor handling your manuscript decide on whether your... is appropriate for that journal. Argue with an editor or reviewer over their decision. If you do object...

...proteins, such as human growth factor, vascular endothelial growth factor, and interleukin-6 (Witkiewicz... was contributed by Bárbara Pinho, a science communicator at the Portuguese science museum "Fábrica Centro...myofibroblasts, neuroendocrine, adipose, immune, inflammatory, angiogenic vascular, endothelial, and stromal... in increased expression of transforming growth factor-β signaling, possibly inducing the secretion of...seems to attack us by releasing mitogenic growth mediators and improving cancer cells proliferation.  How...enzymes and metabolism reprogramming aren’t the whole story. It’s essential for a tumor to spread and to resist... Pinho! Bárbara Pinho is currently a science communicator at the Portuguese science museum "Fábrica Centro...

...is a STEM career counselor, Director of Education for the Jackson Laboratory for Genomic Medicine and a... successful and sustainable biomedical research sector at our first Future of Research symposium. The ...compulsory internship to be taken in an industry, regulatory, or clinical environment. It remains contested...the overall structure of the biomedical research sector, and what a tenable workforce structure may look...ideal and productive workforce look like for the sector – how many jobs are there for STEM graduates and...strong supporter of increasing the number of postdoctoral scientists at University of Maryland. Paula... at Scientific Meetings from Addgene Executive Director Joanne Kamens Learn how to be a successful manager...

... be viral vector jokes and comics. Other new resources for researchers using viral vectors In addition...Addgene blog Download our Viral Vectors 101 eBook Find all of our viral vectors blog posts Learn more about...Starting today, our Twitter feed will be full of viral vector information. Each day, we’ll cover a different ...AddgeneGoesViral to find information on the basics of viral vectors, applications, and some new resources available...also investigates new strategies to improve viral vector production and data analysis. Our first paper came...

...in viral vectors, but easily fit in a TE. Transposons are also less likely than viral vectors to induce...DNA transposons are most commonly used in the laboratory for genome manipulation. When transposons are...transposon’s LTRs, similar to when packaging viral vectors.   Let’s take a closer look at three different ... in human cells at rates similar to retroviral vectors. The hyperactive version of the SB transposase,...efficiency in human cells similar to retroviral vectors. The hyperactive PB transposase (hyPB) is ~10-fold...transfer Transposons are an alternative to viral vector delivery of transgenes, like for iPSC reprogramming... piggyBac, which is a big advantage over viral vectors (~5 kB payload for AAV and ~8 kB payload for lentivirus...

...Hendel et al. 2015). Low detection rates mean the investigator can be relatively certain that their samples...off-target mutations are essential data points for investigators looking to definitively link genotype and phenotype...Targeted amplicon sequencing doesn’t tell the whole story Figure 1: Schematic representations of the...viable options for multiplex sequencing in many laboratories. Nevertheless, unbiased detection isn’t as sensitive...genome-wide unbiased methods. However, most investigators use single cell clones for in vitro CRISPR experiments... Hough. We additionally thank Monica Sentmanat, Victor Dillard, and Ayokunmi Ajetunmobi for their contributions... Hough is a Biochemistry PhD candidate in the laboratory of Prof Steve Jackson at the Gurdon Institute...

...nick, and monitor indel frequencies (Anzalone et al., 2019). Figure 6: Prime editors bound to ... Figure 4: Editing the PAM prevents the prime editor from re-engaging with DNA it has already edited...to pegRNAs drive editing (left), while prime editors bound to sgRNAs only drive nicking (right). Start ...opposite strand. Created with BioRender.com. MMR inhibitors PE4 and PE5: These approaches inhibit MMR to...inhibited. Created with BioRender.com. RNA protectors epegRNAs: epegRNAs include structured motifs...primer binding site (Nelson et al., 2022). PE7: This editor includes the fused RNA-binding protein La to protect...software, however. You may find it useful to ask a mentor what software tools they recommend — feel free ...

...cells survive their own secretions. Pro tip! Historically, colicin-producing plasmids have been referred...referred to as “Col plasmids” or “Col factors.” Virulence plasmids: microbial infection Virulence plasmids...other molecules to host organisms. For instance, vectors derived from the Agrobacterium Ti virulence plasmid... through direct contact. That’s conjugation! F factors can also be integrated into the bacterial chromosome...developing new cloning techniques based on these vectors (Shareck et al., 2004). Plasmid categorization...the many plasmid tools available in the Addgene repository. To learn more about some other common plasmid...Auriol, P., Vogel, T. M., & Simonet, P. (2001). Laboratory-scale evidence for lightning-mediated gene transfer...

...Washington Office Ben Corb - Director of Public Affairs at ASBMB Adam Fagen, PhD - Director, Early Career Scientist..., these issues which will continue to shape the sector throughout their careers (McDowell et al, F1000Research...working group (National Academy of Sciences. “The Postdoctoral Experience Revisited” 2014 Appendix B p93-95...funding and funding mechanisms require an engaged sector with effective advocacy skills. Advocating for ...Scientist Segment at AAAS Marnie Gelbart, PhD - Director of Programs at Personal Genetics Education Project...their respective fields: David Cameron, PhD - Director of Media Communications, Harvard University Ray...Advocate with the Perfect Science Talk Find a Good Mentor ...

...Diagnostics will play an important role in helping to monitor and slow this spread until vaccine programs can... switches for pathogen detection In 2014, as postdoctoral fellows with Jim Collins and Peng Yin at the.... The first was a new class of programmable riboregulators, called toehold switches, which allow for the...allowed gene circuits to be deployed out of the laboratory in a cell-free and biosafe format that was stable...circuit-based sensors to be used outside of the laboratory. The detection threshold for these RNA sequences...the technology through product development, the regulatory process, and manufacturing so that we can get...Leslie Dan Faculty of Pharmacy at the University of Toronto. His research is focused on developing cell-free...

...deliver in low-capacity vectors like AAV. AAV (adeno-associated viral vectors) are commonly used for in...Background Read About CRISPR Competitor NgAgo Learn How You Can Use piggyBac Vectors in Genome Editing Resources...preferential infection of certain tissues. However, these vectors have a low genetic capacity of ~4.5 kb. At 4.1 ...barely fits into AAV when you add in a gRNA and regulatory elements. To solve this problem, the Zhang lab...previously used SaCas9 (1053 aa/3.2 kb) with AAV vectors for efficient postnatal editing. LbCpf1 and AsCpf1...SpCas9. These nucleases should fit well into AAV vectors, especially given that Cpf1 gRNAs are also shorter...

..., 50ms “off”.  A pulse generator such as the A.M.P.I Master-9 Pulse Stimulator, which can control output...cable is threaded through—and fixed to—an optical insulator called a ferrule (below). The fiber optic cable...Channelrhodopsin 2 (Chr2) has been expressed in the motor cortex of a mouse. Upon connection of the ferrule...to give the animal the full range of motion, a commutator allows the patch cable to connect to the laser...University of Michigan and he is currently a Postdoctoral Fellow in the lab of Mary Jeanne Kreek at the... 1. Gutierrez DV, et al. Optogenetic control of motor coordination by Gi/o protein-coupled vertebrate ... neurons. Current protocols in neuroscience / editorial board, Jacqueline N Crawley  [et al]. 2012;Chapter...

... was contributed by Kusumika (Kushi) Mukherjee, editor of Trends in Pharmacological Sciences, a Cell Press...Shinya Yamanaka’s lab, where the reprogramming factors were delivered via retroviral infection [8]. The...Kushi) Mukherjee. Kusumika (Kushi) Mukherjee is the Editor of Trends in Pharmacological Sciences, a Cell Press...communication and publishing after completing her postdoctoral training from Massachusetts General Hospital...PMCID: PMC2752493. 4. Bongso, A. and M. Richards, History and perspective of stem cell research. Best Pract...embryonic and adult fibroblast cultures by defined factors. Cell, 2006. 126(4): p. 663-76. PubMed PMID: 16904174... cells from adult human fibroblasts by defined factors. Cell, 2007. 131(5): p. 861-72. PubMed PMID: 18035408...

...Keep the hood clean Hoods should not be used as storage cabinets. Clutter not only makes it difficult to...air conditioning on) so leave them in a sterile incubator. This also means that any medium or washing buffer...entering the hood and when putting them into the incubator). It also helps to get familiar with the morphology...results. Most cell culture media contain the pH indicator phenol red (pH range 6.0 (yellow) - 8.0 (red))...aliquot them into a number of vials, freeze them, and store all but one in liquid nitrogen. The frozen vials...master stock. This ensures that you have many cells stored in their original condition and can replace your...with the Agarose Method Plasmids 101: Mammalian Vectors Resources on Addgene.org Find mammalian plasmids...

...development programs for bioscience educators. Using these hands-on tools, educators introduce their students to...This post was contributed by Tim Herman, director of the MSOE center for BioMolecular Modeling and the...dangerous. Subscribe to CRISPR Blog Posts How do educators and students use molecular models?   We created...something tangible. Under the guidance of a trained educator, these instances of focused attention become magical... their labs. Soon after we began working with educators in our Center for BioMolecular Modeling, we spun...our guest blogger Tim Herman! TIm Herman is the director of the MSOE center for BioMolecular Modeling and...Addgene.org Visit Our CRISPR Guide Page CRISPR/Cas9 History Browse All CRISPR Plasmids and Resources ...

...guest blogger Melanie Fox, founder and executive director of Central Indiana Science Outreach and a Postdoc...professional development I had a wonderfully supportive mentor and graduate office. I cannot stress how important...my blog helped immensely when CINSO started brainstorming the resources we wanted to provide online. ...in real time! I follow great online science communicators like Phil Plait @BadAstronomer, Emily Graslie...when I was trying to find local informal science educators, and ever since, the State Museum has been the...week. I gained experience explaining science to visitors of all ages, learned a lot about the dinosaurs...Melanie Fox is currently the founder and executive director of Central Indiana Science Outreach and a Postdoc...

...making it easier to construct targeting vectors. Addgene depositor Takashi Yamamoto’s lab has harnessed ...cloned into a vector containing Cas9 and the PITCh-gRNA. Step 3: Contransfect the PITCh vector with the vector...just upstream of a stop codon. Briefly, the PITCh vector should be constructed with 5’ and 3’ microhomology...PITCh Step 1: Generate microhomologies in the PITCh vector ~20 bp 5’ and 3’ microhomologies are added to ..., and this construct is inserted into the PITCh vector via In-Fusion or SLIC cloning. Step 2: Design an... vector carrying Cas9 and the PITCh- and locus-specific gRNAs Step 4: Select for puromycin resistant cells...

...conjunction with activator or repressor domains to control gene expression. Base editors can help you make... in a single lentiviral transfer vector or separate transfer vectors. May contain reporter gene (e.g. ...experiment As with any experiment, there are many factors that need to be considered during the planning ... that would work for your system. Addgene has depositor submitted protocols and links to a CRISPR forum...Components of System Application Mammalian expression vector Promoter driving Cas9 expression can be constitutive...CRISPR elements are inserted into an AAV transfer vector and used to generate AAV particles. Transient or...

...Science describing the development of “transcriptors,” transistor-like digital genetic switches that enable...Stanford develops engineered DNA systems capable of storing data and computing inside living cells. He's also...cellular computing. These logic gates built from transcriptors are available at Addgene as a BIOFAB kit comprised...world using words like OR and AND. These logical operators are so useful they are implemented in most materials...the beginnings of computational logic and data storage. What we don't have are the keyboards and displays...mining. There is an absolutely incredible hidden story here. The parts we are releasing simply plug into... plants. We are really plugging into the "stuff sector" if you will, making chemicals and materials, and...

...signaling pathway with many receptors, signaling molecules, transcription factors, and ligands. The presenter... unbiased discovery. With these screens, the investigator generates a cell population where all genes ...contexts. Most of the immune cells we study are migratory, and their exact localization is important for...types of interactions to a large extent involve receptors expressed on the cell surface. Thus, a screen ...targets, preferentially those for which validated inhibitors or even approved drugs already exist. If you ...The gRNA oligos are subsequently cloned into your vector of choice using the adapter sequences that you ...strategies related to other nucleases as well as inhibitory and activating approaches.  Many thanks to our...