We narrowed to 614 results for: cat.1

...phosphorylated DNA termini to 3 ́-OH DNA termini (1-4). In vitro, ligases (notably T4 DNA ligase) are ... enzymology of DNA repair enzymes.     References 1. Shuman, Stewart, and Christopher D. Lima. "The polynucleotide...thermostable DNA ligase-encoding gene." Gene 109.1 (1991): 1-11. PubMed PMID: 1756968. 7. Lauer, G., et al. "Cloning...or DNA base pair mismatches (8-11). For some applications, there cannot be any end-joining activity, and...regions. Why use a high fidelity ligase? Numerous applications have been developed to take advantage of the...Taq and other thermostable DNA ligases. These applications detect specific nucleotide sequences with high...rounds of melting and annealing allows linear amplification of the probe ligation product. With the right...

...found climbing mountains or playing with his two cats. Keira Havens grew up in Hawaii where she was fascinated...challenges involved in plant synthetic biology. Challenge #1: Public perception of synthetic biology One of the...biotechnologists face is public perception of genetic modification (GM) and synthetic biology. The mistrust and...cannot build what we do not understand, and the complicated and unique metabolic processes of plants are ...

...CRISPR tools.  Distributing and depositing In Figure 1, you can see cumulative numbers of CRISPR plasmid ...new CRISPR depositors each year.    Figure 1: Addgene CRISPR distribution and deposits. (A) Cumulative...estimated scatterplot smoothing. (C) Number of requests per year for pooled libraries, categorized by function...insertion via a chemical modifier. We used these categories to see what kinds of CRISPR tools people were... pooled libraries, although we used different categories (Figure 2C). Note that in Figure 2, the numbers...Cumulative number of deposited CRISPR items, categorized by CRISPR function, by year. (B) Number of requests... years.  Libraries are divided into different categories based on function: knockout (KO), activate, inhibit...

...interrogated by the optical system (Figure 1).   Figure 1: Cells expressing a particular surface ...populations of cells in a complex tissue or sample (1). When combined with cell sorters, this technology... it passes through and scatters in different directions. The degree of scatter is captured by detectors...Light that scatters sideways to the path that it was initially traveling is called side scatter (SSC) and...granulocytes in a plot of forward scatter (FSC) versus side scatter (SSC). Users can gate around their...protein composition. It is used for a variety of applications such as measuring target protein expression ...expression levels, assessing post-translational modifications, determining cell health, analyzing cell cycle stages...

...editors (ABE) mediate A•T-to-G•C base changes (Figure 1), but it can be challenging to make these base changes...dCas to the genomic DNA of interest  Figure 1. When ABEs are in action they bind DNA so that the... from E.coli, named TadA7.10 in ABE7.10)  a catalytically impaired Cas protein (dCas or Cas nickase)  ...accidental insertions and deletions (indel) by the catalytically impaired D10A nickase from ABE7.10 can be a ...the authors created ABE8 constructs using a catalytically “dead” S. pyogenes Cas9 (dC9-ABE8.x-m/d). The...these variants, ABE8 could target many additional locations in the genome. Compared to ABE7.10, NG-ABE8.xm... reduced off-target editing frequencies. For applications that require high DNA editing specificity, for...

...fluorophores are constantly being engineered. Table 1 lists a few common fluorescent proteins and their ...definitions found in Table 2 may not apply. Table 1: The specific mutations comprising common fluorophores...transgenic mice. Purification: GFP can be used as a general epitope tag for protein purification and a number...their experimental needs, and some of the common applications in the lab. Why Green Fluorescent Protein?...to maintain fluorescent activity; very little truncation is tolerated, however, point mutations are acceptable...Re-engineering GFP to increase its range of color and application Almost as soon as its sequence was elucidated...others) Neutral mutations    A multitude of applications Due to its size and ease of use, GFP and other...

...cleavage (Harrington et al., 2017). Figure 1: Anti-CRISPR proteins (Acr's) can block CRISPR activity... ways to anti-CRISPR proteins in your experiments 1. Reduce off-target effects Prolonged Cas activity ..., 2018). For example, AcrIIA4 mentioned above indicates that it’s the fourth Acr protein discovered that...viral particles that have the gene deletion could replicate when challenged with the S. islandicus CRISPR-...B, Bondy-Denomy J (2020) Anti-CRISPR protein applications: natural brakes for CRISPR-Cas technologies....light-inducible Cas9 activity? Check out these these plasmids Catch Up on Your CRISPR Background with Our Guide Pages...

...crevices proteins on the cell surface. Figure 1: Comparison of common antibody, HCab, and nanobody...Humana Press, pp 133–148. https://doi.org/10.1007/978-1-62703-293-3_10 Walter J et al. (2020) Synthetic nanobodies...Starting with libraries and a biotinylated (for purification purposes) protein target of interest, the protocol...-30) of sybody candidates are sequenced for identification. All unique sybodies can be subcloned into ...2020) Generation of synthetic nanobodies against delicate proteins. Nature Protocols 15:1707–1741. https...Light to Manipulate Cell Signaling and Protein Purification RANbodies: Reporter Nanobody Fusions Plasmids...

...Updated Jun 1, 2021 by Meghan Rego. Reproducible data are key to science, so scientists are used to repeating...the DNA packaged inside the viral particles (Figure 1). Briefly, packaged DNA is extracted from purified...These impurities cannot be removed during the purification step because they are inside the virus itself...very low concentration. So how do we make sure to catch all of these potential mishaps before distributing...Guerin et al., 2020).  In addition to genome identification, we developed custom open-access Python scripts...

...used in way more applications.   Fig. 1 Making a Protein Tag (Abridged) (1) Locate a promising monoclonal... by its antibody without additional chemical modification steps and when the epitope was within the tagged...

...Podcast breakdown Intro Chapter 1: Who is Crystal Shih? Crystal's educational background and position at Novartis...Chapter 7: How Crystal developed her science communication skills and how they've helped her at Novartis...

...for example, split cells on a Monday diluting them 1:10, you should be able to split on the following Thursday...Immunology. You can find Sana on LinkedIn.   References 1. Yin, R., et al. "Gene expression profiling analysis...results. Most cell culture media contain the pH indicator phenol red (pH range 6.0 (yellow) - 8.0 (red))...Anything between orange and light orange usually indicates that the cells have grown enough, are making their...

...DYKDDDDK or DYKDDDK 1 Affinity and Purification Good for antibody-based purification; has inherent enterokinase...does not greatly alter the size of the tag.   Table 1: Common protein tags Tag Epitope Mass (kDa) Function...site GST Large Protein 26 Purification and Stability Good for purification with glutathione; protects...used for purification as elution requires low pH poly His HHHHHH  0.8 Affinity and Purification Very small...GKPIPNPLLGLDST   1.4 Affinity and Purification Good for antibody-based purification Combo and cleavage tags...multitude of uses including (but not limited to) purification, detection, solubilization, localization, or...fusion proteins to aid in the expression and purification of recombinant proteins. Tags for stability...

... edit.   Figure 1: Choosing the best PE6 variant for your application. Image reused form Doman...delivery by modular envelope design. Nature Communications, 14(1), 5141. https://doi.org/10.1038/s41467-023... CreVLPs, and eVLPs for in vivo and in vitro applications.   Figure 2: Modular features of DIRECTED...approach for experimental therapies and medical applications.  Find DIRECTED plasmids here! Strebinger, D...interactions or map a local proteome, but some applications have been challenging or inaccessible. The Ting... (which had therefore been trafficked between locations). Figure 3: A) LOV-Turbo becomes active...active when illuminated with blue light, catalyzing the biotinylation of nearby proteins. B) TransitID uses...

...membrane targetin sequence derived either from p63(1-29) and Lck. Image from Chertkova et al., 2020. ...Reports. 2020. https://doi.org/10.1038/s41598-020-57500-1 The CRISPR corner By Jennifer Tsang Here are a few...Fluorescent tools for mammalian cells Nanobody purification CRISPRi and CRISPRa system for hPSCs The CRISPR...CRISPR corner New from the viral service Faster catalysis after yeast-based protein evolution By Joanne ...concept, Sanchez and Ting sought to improve the catalytic efficiency of tobacco etch virus protease (TEV...activity with a fluorescent protein--no affinity purification required! The team used other molecular techniques...evolution platform to generate TEV protease variants. Catalysis by TEV protease allows a transcription factor ...

...Aptamers and Their Biological Applications.” Sensors (Basel). 2012; 12(1): 612–631. PubMed PMID: 22368488...be selected for these small ligands. References 1. Ma, Hanhui, et al. “Multicolor CRISPR labeling of... Aptamer biosensors are used in a variety of applications, including detection of environmental pollutants... opens aptamers for use in many entirely new applications including aptamer based fluorescent biosensors... render this approach less sensitive in most applications. Learn About Fluorescent Biosensors Light-up..., MG proved to be too toxic for many in vivo applications, but this work proved the concept that RNA could...and biochips. Ultimately, there are just some applications that lend themselves to aptamer-based instead...

...base to the other. Figure 1: Base transition editors catalyze purine-to-purine or pyrimidine-to-pyrimidine...engineering and protein delivery. Nature Communications, 8(1), 15790. https://doi.org/10.1038/ncomms15790...SpCas9 base editors with editing windows as small as 1-2 nucleotides. To reduce off-target effects associated...Editors, which we've discussed in two follow-up posts: (1) (2). Base editors have a narrow editing window of...arrows indicate a conversion directly catalyzed by the base editor. Dotted red arrows indicate the change... point modifications. CRISPR base editing persists through multiple cell divisions, indicating that this...target modification in primary T cells, making them a promising tool for cell therapy applications. Dual...

...about health equity, science communication, and public health.     References 1. O'Neill, J. "Tackling drug-resistant... to the development of resistance include (Figure 1): Over-prescription of antibiotics (for example, ...Microbes (bacteria included) are very skilled at the delicate art of survival. Existing virtually ubiquitously...of intentions. The take-home? Always take your medication as prescribed. How do we know which bacteria ...

...Spaghetti Monster probes at Addgene Uses of smFPs 1. Multi-channel connectomic tracers Fluorescent proteins... expanded use of smFPs in the future. References 1. Viswanathan, Sarada, et al. "High-performance probes...controlled by collections of neurons that are intricately wired into circuits through synaptic connections...photostable than GFP. In addition, antibody-based amplification of their signals is weak and increases background...more HA tags. 4. High-resolutions microscopy As indicated above, smFPs can be used with conventional microscopes...

...regulated in the medicinal plant, the Catharanthus roseus (Figure 1). The ultimate goal of our research ...activities in seed plants." Plant cell reports 9.1 (1990): 1-5. PubMed PMID: 24226366. Lee, Min Woo, and Yinong...and a vacuum is applied with a vacuum bell or desiccator. Successful infiltration can often be seen by...expression method for analyzing gene function in Catharanthus roseus seedlings." Frontiers in plant science...

... cerevisiae (Tuller et. al., 2010). Figure 1: An overview of peptide synthesis. A ribosome interacts...McClain WH (2000) The G·U wobble base pair. EMBO Rep 1:18–23 . https://doi.org/10.1093/embo-reports/kvd001...folding and stability, change post-translational modification sites and even affect protein function (Mauro... AP, Bilen B, Zavolan M (2013) Analysis of CDS-located miRNA target sites suggests that they can effectively...synonymous codon usage bias, and its potential applications. Nucl Acids Res 15:1281–1295 . https://doi.org...

... present in the sample (APC in the case of Figure 1). In Figure 1A, the population needs to be shifted... decrease the compensation value.   Figure 1: Compensation example with single color control. Flow...diagonal population, this is a sign of spillover and indicates that it is necessary to apply a compensation value...

...glorious trophy. Sure it’s plastic and you have to share 1 trophy among 6 people, but you’ve earned bragging ...plasmids! Each chose their favorite plasmid and replicated it. Can you guess each plasmid? Our lab paid...creative hijinks last year. They showed us their dedication to great customer support by dressing up as our...

... Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...skipping the preparation steps of production, purification, and titering. Our newest viral vector service...

...a ‘plasmid’. “p” is for plasmid. pXXXXX-XXXX Step 1: Backbone name Include the empty backbone name in ... eukaryotic cells). pBACKBONE-XXXXX Tip: For a catalog of published and commercially available empty backbones...considerations  If your insert contains a mutation or modification, this should be included in the plasmid name...

...package from Addgene.  Here are the details:   Contest 1: Deck the Lab - Classic Edition This edition of the...create a stack of wrapped gifts using life science catalog pages. Make your WFH space feel a little more festive...

...Time for the #BioSci3319 #StreakPlateChallenge part 1. Lab B needs a redo :-( @SandTBioSci #CompetitiveStreaking...blog posts on fluorescent proteins Check out educational resources on the blog Resources on Addgene.org...

... Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...

... necessary for advancing the field. References: 1. Internal initiation of translation of eukaryotic ... of a 2A peptide derived from porcine teschovirus-1 in human cell lines, zebrafish and mice. Kim et al...desktop. The Plasmids 101 eBook is designed to educate all levels of scientists and plasmid lovers and...

...alternative detection methods.      Figure 1: The four types of ELISA are direct, indirect, sandwich...conjugated antibody is not readily available. Table 1: Overview of the advantages and disadvantages of each...staining. Direct ELISAs also lack the signal amplification afforded by secondary antibodies and thus are...

...true reproducibility. The following are critical: (1) connectivity to other data sources used in research...engagement with the Wisconsin research community were to: (1) prevent vendor lock in by making it easy to get data... both accept deposit of data associated with publications into one of five general purpose open source...

...light-sensitive LOV2 domain of Avena Sativa phototropin 1 (AsLOV2). In the dark, this peptide is caged by the...iLID nano and 800nM to 47µM for iLID micro; Figure 1). With this great affinity range, these LIDs have ...Figure 2). They are all available now at Addgene. 1 Lungu et al., Chem Biol. 2012 Apr 20;19(4):507-17....Guntas et al., Proc Natl Acad Sci USA 2015 Jan 6; 112(1):112-7. doi: 10.1073/pnas. CRISPR-Cas9 optogenetic... al., BMC Molecular Biology. 2013 August 20th; 14(1). TRICK: A method for visualizing the first round...for a variety of experimental purposes including (1) single or multiple, constitutive or inducible, gene...the years. The authors optimized these systems by: 1) decreasing the number of plasmids required for each...

...sequence on either side of the desired mutation (usually 1-3 mismatched bases) is sufficient for your primers...researcher at Cornell University.       References 1. Breslauer, Kenneth J., et al. "Predicting DNA duplex...desired modification. Finally, the positive clones are sequenced to confirm the desired modification and ...polymerase activity (for amplification), 3’->5’ exonuclease activity (increases amplification fidelity), and no... genome modifications. Occasionally, multimerization of the PCR primers can cause duplication of the primer...the desired modification have been identified (and confirmed to be free of primer duplications), final validation...and the absense of additional modifications.   Site-directed mutagenesis of Plasmids. A) Generation...

...Figure 1). Clustered events are called a population.                             Figure 1: While...done using side scatter (SSC) and forward scatter (FSC), which measure light that “scatters” past the cell... the plot that divides it into four, using the location of the cell populations as a guide  (Figure 3....be used for this purpose.  Pro tip! The forward scatter is a proxy for diameter and volume of the cell,...the exclusion of doublets and debris. The side scatter roughly measures granularity, a proxy for nucleic... will have numbers by gated populations; this indicates the percentage of total events contained in the... gated population. Heat maps are often used to indicate the relative density of cells. You can see both...

...folding, and deal with sub-optimal codon usage (Table 1) A few mutations are common to all or most expression...curated list of Bacterial Expression Systems Table 1: E. coli expression strains  Note: All strains are...expression vector cannot have p15A origin of replication F- ompT lon hsdSB (rB- mB-) gal dcm (DE3) pLysS...expression vector cannot have p15A origin of replication F- ompT lon hsdSB (rB- mB-) gal dcm (DE3) pLysE...Expression vector cannot have p15A origin of replication F- ompT hsdSB(rB- mB-) gal dcm (DE3) pRARE2 (...Expression vector cannot have p15A origin of replication fhuA2 [lon] ompT gal (λ DE3) [dcm] ∆hsdS/ pLemo...Expression vector cannot have p15A origin of replication F-, Φ80ΔlacM15, thi, lac-, mtl-, recA+, KmR Expression...

...$47,476 and the date for implementation was set for Dec 1 2016. In order to be exempted from the need to pay... these institutions (populations potentially from 1 to 5761, according to the NSF data) and therefore ...Labor’s general guide to the effect on higher education, you can see that those in teaching roles (such...frankly quite surprising. Consortia of higher education institutions submitted letters to the Department...centrally administered, which leads to many complications, including the inability of some institutions...paid. Institutional salary policies are one indicatator of how salaries may changes due to the FLSA. ...of the postdoctoral workforce. Based on our communications with and research on these institutions, we...

...older epitope tags - only time will tell! References 1. Georgieva, Maya V., et al. "Inntags: small self-structured...immunoprecipitation applications, but none of the traditional tags were suitable for both applications. To prove...from non-vertebrate systems. Preliminary testing indicated that inntags IT5, IT6, and IT10 expressed well...and the chaperone Ydj1, both of which display sophisticated interactions with multiple other proteins. Again...detrimental effects on cell size and growth rate, indicating that they compromised protein-protein interactions...These tags are easy to use in multiple common applications. Due to their limited effects on native protein...particularly in single-molecule or high-throughput applications. Perhaps IT5 and IT6 will even replace our older...

...an antibody performed in an experiment.   Figure 1: The front page of the Addgene Data Hub.   Antibody...very little data on which might work in your application. It can be quite difficult to decide on which...be searched and sorted by gene/target, name, application, pass/fail rating, sample species, antibody species... how the antibody performed in the submitted application, along with a brief description of the experiment...experiment. If that data was included in a publication, you'll find that information in the report as well... section will vary slightly depending on the application — flow cytometry, western blot, immunohistochemistry...data showing how the antibody worked in your application. The submission process is easy, with simple,...

...know that most cephalopods die after they lay their 1 clutch of eggs? Propagation is thus quite difficult...organism blog posts: Five Popular Model Organisms, Part 1 Five Popular Model Organisms, Part 2 ... their organismal systems for scientists and educational programs for K-12 students. You can see why I...parenting strategies and make important ecological indicators since species live in water and on land. Current...which have been used to treat heart disease. Caterpillars that eat milkweed generate cardenolides that... poisonous) for bird predators - birds eat a caterpillar once, vomit, and never do it again. Cichlids ... expensive. They are using a cool fluorescent indicator system to rapidly identify recombinants. Arthropods...

...by insertion and deletion frequency. Fig. 1: Overview of TIDE. The input is sequencing trace files...keep your product to knock-in size ratio below 10:1. Small knock-ins Is your knock-in edit small, with...how many copies you are looking for. Secondly, categorize your edit – is it any frameshift mutation? A ...enzyme screening for resistivity to digestion, indicating targeted edits.   Interested in this method...a manageable number of the higher probability locations. If you’re worried about off-target effects, you...Tao Chen, Jos Jonkers, Bas van Steensel. Easy quantification of template-directed CRISPR-Cas9 editing. (...and variant-aware in silico off-target site identification for CRISPR genome editing. (2019) Bioinformatics...

...straight to your materials of interest!   Figure 1: The new search bar.   Pro tip! You can use a manuscript...and easily navigate to educational resources, tools, collections in our catalog, and the Addgene Help ...search bar. And if you’re in the mood to browse our catalog, you can use the links under the search box to ...scrolling up and down the page.  Figure 3: The Education & Tools menu Further down What happens if you ... mission, links to our deposit, request, and educational resources page, and previews for our three most...

...single neurons in the living brain   Figure 1: Plasmid mix to label neuronal morphology (eYFP) and...genOway press release for more information. References 1. Atasoy, D., Y. Aponte, H. H. Su, and S. M. Sternson... gives you a single level of control without modification - either Cre is there or it’s not. Cre-mediated... sites can lead to excision, inversion, and translocation of DNA in a similar fashion as between loxP ...37 °C). Smart molecular evolution led to the identification of FLPe which has a temperature optimum of ...-LDB domains of the fusions allowing Cre/FLP translocate to the nucleus where they exert their recombinase...

...school to a startup and back to academia. Chapter 1: How working at a startup prepared Tom for an academic...foundational research, training, startups, and real applications of SynBio. Chapter 3: Synthetic biology as the...synthetic genome project and constructing ever more complicated biological devices. Chapter 6: - Making research...Addgene Blog Learn about the Synthetic Biology Education Nonprofit BioBuilder Plasmids 101: Golden Gate...

...  Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...

...activity to a transcriptional activation domain (Fig 1). Two ways to use CRISPRa for your research CRISPRa... Stucka R, Hauber J, Lange UC (2016) Targeted HIV-1 Latency Reversal Using CRISPR/Cas9-Derived Transcriptional... (2014) A Protein-Tagging System for Signal Amplification in Gene Expression and Fluorescence Imaging.....org Find Cas9 Activators Find Cas9 Repressors Catch Up on Your CRISPR Background with Our CRISPR Guide...

...breaks (DSBs) or donor DNA templates.  Figure 1: An overview of prime editing. Created with BioRender.com...demonstrated ~20-50% efficiency in HEK293T cells with 1-10% indels. Since then, further innovations have continued...Engineering. https://doi.org/10.1038/s41551-024-01227-1 Yan, J., Oyler-Castrillo, P., Ravisankar, P., Ward... the indicated tool. ~ reflects an innovation that may optionally be combined with the indicated tool....genetic variants have been identified in humans and cataloged in the ClinVar database. Previously developed ...prime editing Less constrained by PAM sequence location The prime editor extends the reach of CRISPR genome...editing typically needs to be optimized for each application, and the efficacy varies widely — it still almost...

...is here to help. Contact us at [email protected] or +1 617-225-9000, or check out our MTA reference pages...results, modifications, and inventions created by the Recipient Right to file patent applications on modifications... about intellectual property rights, pending publications, and commercial use often come up. An MTA can...trials Acknowledgement in recipient’s future publications relating to the material Limited liability...

...Cloning grade DNA Transfection grade DNA Figure 1: Transformation introduces transformation grade plasmids...that have been used during the purification procedure. Good indicators of DNA purity are the absorbance...nm should fall ~1.8 - 2 and give scientists an indication of purity versus protein contaminants. The 260...nm ratios should fall ~2 - 2.2, and provide an indication of purity versus chaotropic agents as guanidine...

... successful after their training.   Figure 1: Professional Development Hub Workshop Attendees and... It's been an honor to work with such a group dedicated to providing the structure and know-how to integrate...example, pd|hub assembled a coalition of higher education organizations committed to advancing career and... inclusive career readiness training in this educational path before we lose more scientists who might...

...’ve come to the right place. Get started with PGC-1 plasmids from Addgene. References: Skeletal muscle...window onto communication between the muscle and brain, which may have therapeutic implications, say Hyo ...