We narrowed to 1,042 results for: URE

...office or other authorized office directly to make sure the deposit agreement/MTA is taken care of and, ...Deposit” and “Download Deposit Spreadsheet” (see Figure 1). Please read our “How to Deposit Your Plasmids...thorough guide to our online data entry process. Figure 1: Begin the deposit process by clicking “Submit...Required columns are shown in red in the spreadsheet (Figure 2) and are described in Table 1. Some cells have...Growth Temp Choose from 30 °C, 37 °C, or room temperature. Growth Strain Please indicate which growth...Filling out and sending the deposit spreadsheet Figure 2: Example of the deposit spreadsheet showing the...update the spreadsheet every so often and this ensures you are using our most up-to-date version! We hope...

...we’re really borrowing these tools from Mother Nature’s toolbox. Natural plasmids, also called environmental...the basis of many molecular biology tools! Figure 1: Common functions of natural plasmids. Created... to survive where others can’t. This ability to endure a harsh environment is distinct from the benefit...new types of plasmids without clear analogues in nature. For example, helper plasmids were developed to... To learn more about some other common plasmid features, such as origins of replication and promoters,... blog as scientists continue to explore Mother Nature’s toolbox for new and exciting molecular technologies...Resources References Demanèche, S., Bertolla, F., Buret, F., Nalin, R., Sailland, A., Auriol, P., Vogel,...

...focus was on editing animal models of disease and cultured cells. For example, in 2013, Jinsong Li and colleagues...When all was said and done, the researchers had cured 24 mice of their disease. Meanwhile, Hans Clevers...significant progress in clinical trials.     Figure 1: Types of delivery and editing strategies for...success rate in clearing misfolded protein levels as measured in the blood. More information is expected soon...blindness, and diabetes. Directions and obstacles for future CRISPR therapies CRISPR technology is rapidly evolving...strides since we first wrote this blog post. The future success of CRISPR in the clinical will rely on ...MA, Berkhout B, Herrera-Carrillo E. A CRISPR-Cas Cure for HIV/AIDS. Int J Mol Sci. 2023 Jan 13;24(2):1563...

...Did you say… easier? Fragmid’s modular assembly structure takes a lot of the design work out of the plasmid...Fragmid components on the Fragmid website.   Figure 1: (Top) Schematic overview and timeline of the...schematized. Sizes of fragments are not to scale. Figure and caption from McGee et al., 2024, used under...designing a CRISPR vector the Fragmid way!    Figure 1: Selecting your vector type on the Fragmid website...website.   In step one, you’ll select the features of your vector, which are: Organism (Mammalian...mechanism (KO, CRISPRa, CRISPRi, editing)   Figure 2: Selecting module fragments on the Fragmid website...want? You can make your own Fragmid components! Ensure your backbone and genes contain only the Bbsl cut...

...compatibility and high DNA specificity." Nature (2018) doi: 10.1038/nature26155 PubMed PMID: 29512652 Plasmids...loci or genomes. Cas9s from other species, like S. aureus and N. meningitidis require different PAM sites...first restored catalytic residues D10 and H840. To ensure that the clones were capable of cleavage, they ...fused to third generation cytidine base editor architecture (BE3) slightly improves NGG PAM editing efficiency...efficiencies (edit frequency in cell population measured with NGS) are shown as percentages, with fold ...lot we still don’t know about xCas9. “I’m not 100% sure xCas9 is going to be flat out better than SpCas9...

...the CRISPR/Cas9 system. Nature 516:423–427 . https://doi.org/10.1038/nature13902 Maggio I, Holkers M, Liu... the target cells (Beatty et al., 2012). These features make AdV an attractive candidate for in vivo delivery...genome editing machinery such as CRISPR-Cas. Figure 1: Schematic of targeted gene knock-out (left) ...Gonçalves' lab found in a second paper published in Nature Methods the same year that delivering RNA-guided...double-stranded AdV genomes. Such protein-DNA structures presumably reduce the likelihood that donor DNA... that advantages of AdVs include their episomal nature and very efficient introduction of DNA into therapeutically...

...sequencing (NGS) to identify genes of interest. Figure 1: Overview of in vitro AAV CRISPR screens. Created... capsids, or serotypes, have been identified in nature, while many more have been engineered in labs. ...improve infection rates in your experiment. Figure 2: Visual representation of common AAV serotypes...smaller Cas9 ortholog like SaCas9. You could even venture away from the comfort of Cas9, and opt for a completely...design and execution of CRISPR in vivo screens. Nature Protocols, 17(9), 1903–1925. https://doi.org/10.1038...vector integration into CRISPR-induced DNA breaks. Nature Communications, 10(1). https://doi.org/10.1038/... linkage analysis with in vivo AAV-Perturb-seq. Nature, 622(7982), 367–375. https://doi.org/10.1038/s41586...

...electrical potential. Thankfully, there is a tool to measure these electrical events! Voltage indicators are...They can be targeted to the plasma membrane to measure membrane potential and can also be directed to ...applications such as in vivo tissue imaging.   Figure 1: Ace-mScarlet is a fusion between the voltage-sensitive...depending on your experimental system.   Figure 2: General overview of chemigenetic tools. A genetically...vitro and a small molecule is added exogenously to measure a phenotypic response in the form of fluorescence...Indicators As previously mentioned, cellular voltage is measured at the cell membrane, and this voltage is particularly...has you covered. Happy voltage sensing!   Figure 3: Components of the types of voltage indicators...

... to quickly check out the plasmid's structural features and see if I can easily clone my gene into it?...plasmid map based off of the plasmid sequence and feature information given by our depositors. Many depositors...plasmids are sent as bacterial stabs at room temperature. Core of the plasmid page - Core of the plasmid...), antibiotic resistance, and optimal growth temperature are listed, as well as the name of the strain... Citing this Plasmid: This is a relatively new feature of our plasmid pages, and provides a list of publications...our blog post) for more information about this feature!).  This means you can see which labs have already...the Addgene Blog Read about Our New Citations Feature Check out Our Walkthrough of the Deposit Process...

...exclusively to in vitro cell culture. The user should review the current literature about their preferred model...single-stranded DNA oligonucleotide, so you will need to make sure your expression system and delivery methods are ...system, a good first step would be to consult the literature to see if anyone has published a protocol that...in in vitro transcription reactions to generate mature Cas9 mRNA and gRNA. RNA is delivered to target ...different methods, so we've summarized some key features of common  delivery methods below: Chemical transfection...Resources on the Addgene Blog Check out our CRISPR Featured Topic Page Learn about CRISPR Delivery Using Ribonucleoproteins...

.... Figure 1: Like this jar of jellybeans, a pooled CRISPR library is a complex mixture. Pooled libraries...experimental system are absolutely necessary to ensure that you select the correct CRISPR library. Choosing...concentration that is sufficient to generate lentivirus. Be sure to use next-generation sequencing to check the quality...cells gain resistance to drugs and can identify future therapeutic targets for diseases causing uncontrolled...uncontrolled cell growth, such as cancer. Figure 2: The CRISPR library must be amplified (A) to be ...the screen should be independently validated to ensure that the desired modification produces the phenotype...

...re tethered to bring them into close proximity (Figure 1). Formation of the complete FP results in a fluorescent...can be detected by fluorescence microscopy. Figure 1: How BiFC works. A split FP (e.g., yellow fluorescent...localization of the studied PIPs. The straightforward nature of BiFC also makes it easier to generate a large...fragments and, preferably, some information about the structures of the tested PIPs. Many split FPs have been ...BiFC experiments only require that you image and measure the fluorescence of a single FP. Your control construct...one another — which means it cannot be used to measure the dissociation of PIPs. Thus, whereas BiFC is...fluorescence complementation (BiFC): A 5-year update and future perspectives. Biotech 53. https://doi.org/10.2144...

...causing a lack of exposure, a lack of immunity, and then finally a synchronized exposure under these conditions...conditions once the measures were lifted (Servellita et al., 2023).  This anecdote goes toward saying that...certain contexts (Bauzon et al., 2003).        Figure 1: The adenovirus life cycle. Image reused from...31749409. Harrach, B. (2014). Adenoviruses: General Features. In Reference Module in Biomedical Sciences. Elsevier... Undergoing Spontaneous Degeneration in Tissue Culture. Proc Soc Exp Biol Med. 84(3), 570–573. https:/...type 2 in US children with acute severe hepatitis. Nature, 617(7961), 574–580. https://doi.org/10.1038/s41586...

...center of the structure, shielded from quenching by aqueous solvent. This tightly-packed structure explains...Fluorescent Protein (GFP) has likely existed in creatures such as jellyfish for millions of years; however...while still maintaining fluorescence. The protein structure, first reported in 1996, is an eleven β-sheet-...yield for BFP Y145A and H148D Stabilizes the structure of Cerulean derivatives F99S, M153T, V163A Improves...Optogenetics: Scientists can use light to detect, measure, and control molecular signals, cells, and groups...This is a type of flow cytometry that separates mixtures of cells into distinct populations based on fluorescent...

...Kwapata now works as a lecturer in molecular genetics and biotechnology for Horticulture and Forest Crops at... at the Lilongwe University of Agriculture and Natural Resources. He recently received an award from nonprofit...Kingdom ended up working in the life sciences and agriculture 5:47-9:35 : Kingdom's biotechnology research ...transition back to Malawi and Lilongwe University of Agriculture and Natural Resources 10:30-11:41 : Kingdom on...Seeding Lab's Work with Lilongwe Univeristy of Agriculture and Natural Resources Watch Dr. Kwapata's Speech...

...exploring nature in the real world. Additional Resources on the Addgene Blog Browse our CRISPR featured topic...frameshift or 21+ bp indel. This score is a useful measure to determine how many of the contributing indels...CRISPR-Cas9 double-strand DNA breaks using SpCas9. More features will be added to the ICE software tool soon. To... To request other nucleases or other features and functionalities, please email [email protected]. ...sequence for the nuclease used. Currently, ICE is configured for Cas9 fromStreptococcus pyogenes (SpCas9)....frameshift or 21+ bp indel. This score is a useful measure for those who are interested in understanding how...

...externally fertilized, transparent progeny (Figure 2). These features give it a leg up (or a fin up?) over other...become such a popular model organism? Zebrafish (Figure 1) were initially recognized as a useful vertebrate... tracing in zebrafish To highlight some of the features that make zebrafish useful, I’ll outline two recent...of foreign DNA. Despite these caveats, several features of zebrafish—including the available genetic resources...compensation triggered by mutant mRNA degradation." Nature 568.7751 (2019): 193. PubMed PMID: 30944477. Farrell...cellular resolution in freely swimming zebrafish." Nature methods 14.11 (2017): 1107. PubMed PMID: 28892088...

... there. Make sure you read the instructions for whatever antibiotic you choose to ensure it works in your...shared by every living thing fit onto a single lecture slide. That shared language is the basis of shuttle...are designed to shuttle between species. Figure 1: Shuttle vectors are vectors that can function... your vector between, you should have a plan to ensure it persists in both contexts. For a vector ...This is useful for keeping your eukaryotic cell cultures bacteria-free, but it won’t allow you to select... producer cells, which have the machinery to manufacture infectious viral particles. Those particles are...

...transducing your virus. Let’s start there!     Figure 1: Flowchart showing the process for using pooled...coli are allowed to grow on plates or in liquid culture to amplify the library, followed by DNA isolation... sequencing and further data analysis. Based on Figure 31.5.2 from https://pubmed.ncbi.nlm.nih.gov/27366891... and replicate, then purify the DNA.     Figure 2: A pooled library page with protocols boxed in...is controlled by viral titer, so you’ll need to measure your viral titer before you can transduce your ...with titer, cell type, and other factors, so make sure to put that aliquot to full use! Common conditions...that a cell doesn’t get infected at all. To make sure that every plasmid’s payload is adequately represented...

... host cell's initiation proteins. In nature, such plasmids ensure their propagation by limiting their ...replication in bacteria is carefully regulated to ensure that it occurs at the appropriate time—for example...lower levels of DnaA to get the job done.   Figure 1: DNA melting by DnaA and IHF is the first step...Termination sequences present in the chromosomal DNA ensure that the helicases are released from the DNA once...of the chromosome, ending replication.   Figure 2: Prokaryotic chromosomal replication. Adapted...high, restraining further replication.   Figure 3: Comparison of replication regulation mechanisms...

...for at least 12 hours, or at lower temperatures for longer. Cultures induced with arabinose and colonies...results in the production of air-filled protein nanostructures called gas vesicles, which scatter sound waves..., used in bacteria, expressed poorly at body temperature (37oC) and were too burdensome to express in ...Excess gvpA plasmid added into the transfection mixture in HEK293T cells gave drastic improvement in expression...transfection reagent (e.g. PEI-MAX). Simply cotransfect a mixture of 1248 ng of CMV-gvpA-IRES-BFP plasmid and 954...noninvasive imaging of microorganisms in mammalian hosts. Nature 553, 86–90 (2018). Farhadi, A., Ho, G. H., Sawyer...

...or resume). If you receive a card in return, make sure to follow up with a professional “thank you” via...your CV doesn’t necessarily mean that you are a future employee of the company. It does, however, offer...to consider, some of the best options are the Naturejobs Career Expo and the ACS career fair, which are... candidates. If you do receive such a call, make sure to get that person’s contact information and again...when you are looking to change positions in the future. Anyone looking for work in today’s job market ...with you to ease the way. Make connections, and nurture those relationships. Then when you are ready to...job, you’ll be in the best possible position to secure it! Further Reading and Outside Resources http...

...engineered CRISPR-Cas9 complex. Nature 517:583–588 . https://doi.org/10.1038/nature14136 Pan J, Meyers RM, Michel...screens can be used to gain insight into protein structure and function (Pan et al., 2018). cDNA libraries... therapeutic targets using CRISPR–Cas9 screens. Nature 568:511–516 . https://doi.org/10.1038/s41586-019...C, Hsu PD, Habib N, Gootenberg JS, Nishimasu H, Nureki O, Zhang F (2014) Genome-scale transcriptional ...2018) Interrogation of Mammalian Protein Complex Structure, Function, and Membership Using Genome-Scale Fitness...Zetsche B, Volz SE, Zhang F (2015) A split-Cas9 architecture for inducible genome editing and transcription...

...datasets with their resultant figures. Instead of providing a “dead end” figure or table, clicking on the ... descriptions, legends and limiting numbers of figures such that many have multiple, complex panels. PLOS...sizes) but images should be unprocessed and in capture format. This raises another issue of proprietary...Other new data initiatives are also taking root. Nature Publishing Group recently launched a publication... and funders. Clearly, it is in our interest to ensure that our data are reproducible and useful to others... alpha isoform of GSK-3 to the same (deserved) stature as the beta isoform. Follow him on Twitter @jwoodgett...

... biggest change is to our most frequently used feature: the search bar. We’ve changed the colors and centered...to go straight to your materials of interest!   Figure 1: The new search bar.   Pro tip! You can use a...will prompt you to log in or create an account.  Figure 2: The new deposit materials bar New menu Most ...website without scrolling up and down the page.  Figure 3: The Education & Tools menu Further down What...look and a fresh new friend: Antibody of the Day!  Figure 4: The plasmid, viral prep, and antibody of the...t take our word for it — check out all the new features and designs addgene.org! And when you’re finished...

... "Inntags: small self-structured epitopes for innocuous protein tagging." Nature methods (2015). PubMed...each tagged protein must be tested carefully to ensure it retains the attributes of the native protein...Georgieva et al. to develop new tags with defined 3D structures less likely to compromise protein folding and...Due to their limited effects on native protein structure, inntags may become the tags of choice for examining...correlation for protein localization in mammalian cells." Nature methods 10.4 (2013): 315-323. Pubmed PMID: 23435261...

...Addgene’s repository for use in neurons, both in culture and in living brain tissue. “The brain is such ... that’s exactly what the LITEs now enable. In a Nature paper introducing the optogenetic tool, Konermann...said in MIT’s press release. In an accompanying Nature News & Views Andreas Möglich and Peter Hegemann...of transcription occurs within minutes of light exposure. Third, the response can be graded with light ...applied non-invasively, its use is not restricted to cultured cells but extends to freely moving animals, as...endogenous transcription and epigenetic states." Nature. 500, 472–476 (22 August 2013).   , with the ...

... each requires selection. These two steps are (Figure 1): The initial plasmid integration step (the ...antibiotic resistance marker from the vector (abR, Figure 1). The resolution step (“double crossover”) which...for the loss of a specific phenotype (using sacB, Figure 1, further discussed below).  Counter-selectable...frequently requires considerable optimization to ensure strong counter-selection. Anecdotally, there is... of two visible blue or magenta chromoproteins (Figure 2) (Liljeruhm et al., 2018). This system is a nice...aeruginosa genome with two-step allelic exchange." Nature protocols 10.11 (2015): 1820. PubMed PMID: 26492139...of bacterial genomes using CRISPR-Cas systems." Nature biotechnology31.3 (2013): 233. PubMed PMID: 23360965...

...mammalian evolution. Nature 428, 493–521 (2004). https://doi.org/10.1038/nature02426 Iannaccone, P. M., ...processes and developments. Model organisms often feature genetic, physiological, or developmental characteristics...laboratory settings. One of its most attractive features is its stability and easily obtainable growing...axolotl has the ability to regenerate complex structures such as limbs, the spinal cord, and internal ...disease research. One of their most attractive features are their eggs. Frog eggs are large, opaque, and...genomics study in bacteria with superior performance. Nature Communications, 9(1). https://doi.org/10.1038/s41467...

...get several interviews but no job offers, is a failure to clearly articulate one's talents and ideas. ...Have a competitive CV You don't necessarily need a Nature/Cell/Science paper. You do need multiple first/...techniques. You DO need to be prepared to know the literature of your field and all of the caveats and alternatives...misfolded proteins is essential for human health. Failure to do so is implicated in several human diseases...much easier to focus on the questions when you’re secure in the knowledge that you have given everyone a...you want to convey for each section or aim. Make sure those get priority. The fine details for aficionados... is the task. Do your homework. Learn what your future potential colleagues study. Think about how you...