We narrowed to 657 results for: cat.2

...positive cells (Martin et al., 2019). Figure 2: Editing efficiencies using base editing reporters...2019). TREE is a real-time, fluorescent based identification system for the isolation of base-edited cell...to BFP Conversion: A Versatile Assay for the Quantification of CRISPR/Cas9-mediated Genome Editing. Molecular...Harris RS (2018) A fluorescent reporter for quantification and enrichment of DNA editing by APOBEC–Cas9...

...phosphate monobasic monohydrate (NaH 2 PO 4 ∙H 2 O), pH 7.0 138 g NaH 2 PO 4 ∙H 2 O 1 L deionized water Adjust...sodium phosphate dibasic (NaH 2 PO 4 ), pH 7.0 142 g of NaH 2 PO 4 ∙H 2 O 1 L deionized water Adjust pH...monobasic monohydrate (NaH 2 PO 4 ∙H 2 O) 610 mL of sterile sodium phosphate dibasic (NaH 2 PO 4 ) Store up to...recombinant antibody. Section 2: Buffer exchange Choose Option 1 or Option 2 based on the concentration ...antibodies. Workflow Timeline Day 1: Purify antibody Day 2 or later: Buffer exchange Equipment Class II, Type...conical tubes NanoDrop spectrophotometer 37 °C, 5% CO 2 incubator with shaking platform set to 120 rpm 37 ... channel pipette 20–200 µL single channel pipette 2–20 µL single channel pipette Reagents Aspirating pipette...

...start selecting and testing systemic capsids! Figure 2 contains a decision tree for selecting a capsid, while... highlighted below the table.     Figure 2: This decision tree can help guide your choice on ...transfer to the adult brain. Nature Biotechnology, 34(2), 204–209. https://doi.org/10.1038/nbt.3440 Goertsen... 2023 * indicates viral preps available. Mechanisms of crossing the BBB are indicated in parentheses...suitability of a systemic capsid for an individual application is dependent on a number of factors, including...test your chosen capsid before using it in an application.     Figure 1: AAV toolkit for targeted...doses are recommended for all systemic capsid applications. For example, AAV-PHP.eB is typically administered...

...as the protocol remains the same.    Figure 2: Microscopy-based antibody experiments should include...results.  Replicates Replicates are another critical control in biological experiments. Replicates can be...of biological replicates such as multiple mice in each group and C, technical replicates or repeated measurements...secondary antibodies.   Application-specific controls Some antibody-based applications have very specific ...then moving on to controls for antibody-related applications. After reading, you will have a better understanding...blot and look for the molecular weight shifts indicative of tau phosphorylation. By including the positive...statistical power of an experiment. Technical replicates are repeated measurements of the same sample ...

... more from the authors in this blog post. Figure 2: iGluSnFR3 displays enhanced brightness, expression...sensors One of the latest additions to Addgene’s catalog is a new class of sensors for neuropeptides and...doi: 10.1038/s41592-022-01597-x. New glutamate indicator iGluSnFR3 Improving on the glutamate sensor iGluSnFR...They generated variants optimized for different applications (v82 has higher affinity for glutamate but slower...PDGFR.codonopt    Aggarwal A, et al. Glutamate indicators with improved activation kinetics and localization...have developed a new genetically-encoded voltage indicator (GEVI) called JEDI-2P (Liu et al. 2022) optimized...Sustained deep-tissue voltage recording using a fast indicator evolved for two-photon microscopy. Cell. 2022 ...

... iRFP713, % Oligomeric state Photo-stabilityc, t1/2, s pKa Brightness in HeLa cells vs. iRFP713, % d ...calcium indicator (Shemetov et al., 2021). In addition to FRET, intensiometric calcium indicators NIR-GECOs...characteristics, performance, and applications of near-infrared calcium indicators in this recent review (Shcherbakova...information on NIR FPs, their properties and applications. At the end, there is a Table that summarizes...photoreceptors incorporating biliverdin IXα (BV) autocatalytically. Fortunately for probe developers and users...vivo probes, due to low autofluorescence, low scattering, and deep penetration of red-shifted light. The...their enhanced versions named emiRFPs in your applications.  There is a whole set of spectrally distinct...

...Nutr, 129 (2009a): 477–484. PubMed PMID: 10064313. 2. Chapman-Smith, A., & Cronan, J. E. “The enzymatic...molecular biology applications for biotin and avidin. A short list of some other common applications and links...downstream application. Biotin labelled oligonucleotides can be used in a multitude of applications including... carboxylase and decarboxylases. These enzymes catalyze the transfer of carboxyl groups to organic acids...this highly specific post-translational protein modification. Most organisms have less than five enzymes ...uses of biotinylation in the lab are protein purification and immunoprecipitation experiments. Biotinylated...sites found on avidin are also ideal for signal amplification during these experiments. During detection, ...

...become a valuable asset yourself.     Fig 2: A diagram of the Thomas-Kilmann Conflict Model, courtesy...conflict is not always what was communicated but how it was communicated. The next step is to come up with...of lapsed communication. Making some assumptions about how something should be communicated is necessary...skill in and of itself. Conflict can also be categorized as either “personal” or “substantive”. A personality-based...traced back to one of a few trouble spots:  Communication (or lack thereof): Information sharing gets ...organization gets, and creating new avenues of communication can create as many problems as it can solve....meeting? Different teams frequently prefer to communicate in different ways, so the source of the conflict...

...two visible blue or magenta chromoproteins (Figure 2) (Liljeruhm et al., 2018). This system is a nice confirmation...resistance cassette, and the R6K origin of replication to allow replication only in hosts carrying the bacteriophage...chromosome. These methods roughly fall into two categories: those that rely on exogenous enzymes and those...Lambda Red recombineering, which uses enzymes domesticated from the Lambda bacteriophage, is arguably the...enzymes can also accomplish the desired genomic modification. This technique is frequently called allelic...recipient of interest. Because pDS132 cannot be replicated in the recipient, the only antibiotic resistant...exchange. To do this, we performed the following modifications to pDS132 (Philippe et al., 2004): Replaced...

...excise the HIV-1 genome from every infected cell and 2. prevent reinfection of those cells. Kaminski et al...pre-treatment levels due to the viral reservoirs located in latently infected cells. Although ART-treated...toxic proteins, CRISPR/Cas9 SAM caused apoptosis, indicating a true “shock and kill” response. Cutting HIV...create mutations in genes essential to HIV-1 replication. However, this approach could permit some viral...along with Cas9 in the T-cell line 2D10. PCR amplification and Sanger sequencing of pooled samples showed... HIV-1 could escape from CRISPR/Cas9-induced modification targeting either the LTRs or essential genes...genes. Many of these escape mutations were located near the Cas9 cut site, leading Wang et al. to conclude...

..., whereas the mVenus-APT1 fusion does not (figure 2). In the mVenus-APT1, a lipidation motif, which is...study its function is one of the most popular applications of fluorescent proteins. These fusion proteins...steric hindrance. For instance, GFP may occlude catalytic sites or obstruct binding sites or motifs that...that are necessary for post-translational modifications. Despite these potential issues, GFP has been used...linker sequence short In the early days of GFP applications, many were concerned with steric hindrance by...N-terminus of the donor fluorescent protein can be truncated to increase the FRET efficiency. Creating fusions...fluorescent protein and protein of interest (POI). To indicate the sequence, people often use terminology like...

... Methods 2009; 6, 343-345. PubMed PMID: 19363495. 2. Wang JW, Wang A, Li K, Wang B, Jin S, Reiser M, Lockey...neighboring fragments can be created via PCR amplification with primers that contain the appropriate homologous... tools that help you design primers for PCR amplification of fragments to incorporate such regions of ...assembly meets CRISPR Gibson can be adapted to more complicated cloning schemes, such as those where the vector...CRISPR technology and is described in a recent publication by the Lockey lab (Wang, et al. 2015). In this...aspects of the SLIC methods. First, it uses a dedicated 5’ exonuclease instead of using the exonuclease...DNA fragments contain these sites at the correct location and NOT in the middle of your insert. Gateway ...

... Figure 2: Mechanisms of deaminase-based adenine transversion editors. Red arrows indicate a conversion...conversion directly catalyzed by the base editor. Black arrows indicate natural cellular processes. Created...researchers can now select the best tool for their application from among many options.  Conclusion With these...engineered human uracil DNA glycosylase. Nature Communications, 15(1), 4897. https://doi.org/10.1038/s41467...via engineered uracil-DNA glycosylase. Nature Communications, 15(1), 6397. https://doi.org/10.1038/s41467...

...15.4%) 46 (59%) of these students were in the last 2 years of their degree. Students filled in a pre-program...Universities from five different geographical locations in Spain were enrolled: the University of Valencia...2014/2015, took place on July 27th, 2015 at a dedicated international mentoring conference held at the... own part, mentors receive immense personal gratification as they witness their student’s progress and...like IMP build bridges and global networks. They catalyze the development of knowledge, experience, resources...Mentoring Foundation for the Advancemnet of Higher Education website Resources at Addgene Read Other Career...

...variety of environmental conditions, and double every 2 hours. Yeast are also the first eukaryotic genome ... throughout their lifetime. Thus any genetic modification, such as expression of fluorescent protein or...studying somatic mutations and cancer." Nature Education 1.1 (2008): 183. Zuber, Johannes, et al. "Mouse...

... column gradient for AAV purification. Workflow Timeline Day 1: Purify Day 2: Buffer exchange and concentration...) . Figure 2: Left panel: Iodixanol gradient after ultracentrifugation. The arrow indicates the 60–40%...7.4 1X PBS-MK buffer 100X Poloxamer 188 NaCl MgCl 2 KCl Centrifugal filter units (MWCO 100 kDa) Reagent...PBS-MK buffer Dissolve 5.84 g of NaCl, 26.3 mg of MgCl 2 and 14.91 mg of KCl in 1× PBS in a final volume of...at 4 °C. 1X PBS-MK buffer Dissolve 26.3 mg of MgCl 2 , and 14.91 mg of KCl in 1× PBS in a final volume ...(C) (formulation buffer) Add 5 mL of Buffer B and 2 mL of 5 M NaCl to 43 mL PBS Procedure Preparation ...need more time, you can alternatively centrifuge for 2 h at 200,000 x g at 18 °C. Carefully take the QuickSeal...

...Figure 2: A) When proteins (blue) interact with the air-water interface (AWI) before vitrification, they...beads through an assembly of biotinylated SpyTag-SpyCatcher linkers and spacer proteins ending with a GFP... (Figure 1). This enrichment streamlines the purification and minimizes sample loss while also supporting...scFvs for other targets or could be used for applications beyond cryo-EM.   Find plasmids for MagIC-Cryo-EM...chemical crosslinking or tilted-stage imaging. A truncated form of a tardigrade LEA, RvLEAMshort, protected...optimizing prime editing systems for specific applications! Delivering prime editors (PEs) via engineered...be delivered in a single package. In a recent publication, the Liu lab showed that this method of delivery...

...improvement in muscle physiological performance.  2. Trans-splicing Overview: For trans-plicing, splice...that the oversized AAV genomes are fragmented or truncated when packaged and then following transduction,...and may need to be optimized for each unique application (Trapani et al., 2015).  Expression of smaller...transgene to help prevent the accumulation of such truncated proteins. Examples of use:  Ghosh et al., 2008...design, donor A may be capable of expressing a truncated protein. This can be avoided by excluding a stop...JS (2011) Gene Therapy of mdx Mice With Large Truncated Dystrophins Generated by Recombination Using rAAV6...Improved dual AAV vectors with reduced expression of truncated proteins are safe and effective in the retina ...

...signal separately for each target.   Figure 2: Examples of simultaneous multiplex IF labeling of...protein is (localization) and how much there is (quantification). But what if you’ve got several interesting...primary antibodies that have been validated in IF applications (find IHC validated or ICC validated antibodies...informative image-based figures for scientific publications. PLOS Biology, 19(3), e3001161. https://doi....Immunocytochemistry Protocols for other antibody applications...

... 10 90 2 x 10 8 10 of 2 x 10 8 dilution 90 2 x 10 7 10 of 2 x 10 7 dilution 90 2 x 10 6 10 of 2 x 10 6...6 dilution 90 2 x 10 5 10 of 2 x 10 5 dilution 90 2 x 10 4 10 of 2 x 10 4 dilution 90 2 x 10 3 Pro-Tip...dilutions, in duplicate, of your standard curve plasmid (2 x 10 9 stock made in step #1): Volume of 2 x 10 9 ...molecules/μL To obtain a solution at 2 x 10 9 molecules/μL: 1.59 x 10 11 / 2 x 10 9 = 79.8X dilution ...stock 45 uL 10X 10X Dilution 2 5 uL Dil. 1 95 uL 20X 200X Dilution 3 20 uL Dil. 2 80 uL 5X 1000X Dilution ...valid 8-point standard curve. Figure 2: Example of the amplification plots obtained from an AAV sample. ...PCR for the detection and quantification of adeno-associated virus serotype 2-derived inverted terminal...