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...editors for more details on how they work!) Figure 1: Base transitions are edits between the purine...side groups of DNA bases and leave the core ring structure intact, allowing them to make 4 out of the possible... Cas9 are sequence specificity and the R-loop structure. In order to bind DNA, Cas9 requires (1) a guide... a region of separate strands called an R-loop (Figure 2). This allows the guide RNA to base pair with...accessible on the outside of the enzyme. Figure 2: CRISPR R-loop. Created with BioRender.com. ...abasic sites with A's, producing C → A edits. Figure 3: Mechanism of cytosine transversion editors. ...experimented with adding different proteins to the CGBE architecture. They conducted a CRISPRi screen to identify...

...kinetics which are illustrated in figures 2 and 3.  Figure 2: Glutamate titration curves for ...from Aggarwal et al…, Supplementary Figure 3.   Figure 3: pH titration curves and ∆F/F0 for...recommend using the SGZ for in-culture work only, GPI anchors for in-culture work and 2P in vivo imaging,...using both 1-photon and 2-photon imaging.   Figure 1: 1P and 2P excitation and emission spectra of...iGluSnFR3 v857. Image adapted from Aggarwal et al., Figure 1   Increased versatility through two different...Image adapted from Aggarwal et al…, Supplementary Figure 2.   Improved trafficking and localization... constructs. Image adapted from Aggarwal et al… Figure 4. PDGFR is a C-terminal fusion to the PDGFR transmembrane...

...content we store in them: music, emails, chats, pictures, and apps. Throughout my time as a graduate student...allowing you to upload anything from protocols to pictures of gels or even sketches to your notebook. The...saving trees! The Earth thanks you. 2. Gibco Cell Culture by Gibco Whenever I start to learn a new technique...I digress. When I started learning proper cell culture technique, I tried to find an app that could help...calculations. This is how I came across the Gibco Cell Culture app. Gibco organizes its app into four categories...are a lifesaver when your research requires cell culture and include a counter, lab timer, dilution calculator...calculator, and a couple of other nice features. This app also has a great educational section. This section...

... (see Figure 1). Oligo 2: Add 'ggcc' to the 5′ end of your reverse complement guide (see Figure 1). Perform... tissue culture-free heritable genome editing using RNA virus-based delivery in wheat. Nature Plants. ...single guide RNA (sgRNA) used in Cas systems. The structure is also reminiscent of sgRNAs, with reRNA containing...gene-edited plants without the need for tissue culture (Maher et al., 2020; Qiao et al., 2025). However...choice of promoter for ωRNA (reRNA) expression (see Figure 1). Both plasmids contain a hygromycin resistance...phosphorylation and annealing of the two oligos. Figure 1: Components of TnpB, schematic of pKb-TnpB1 and...transformation into plant cells. Conclusion & Future Directions TnpB represents a new generation of ...

... the process. Also refer to Figure 1 for a visual overview! Figure 1: General overview of the MoClo...dive directly into the pipetting! MoClo nomenclature and structure can be quite confusing for new users,...insert-to-vector ratio using the equation in Figure 3.    Figure 3: Calculation for the plasmid volume... MoClo a walk in the park.  Before you begin, be sure to check out Addgene’s MoClo guide for all the basics...assembly process to produce a multigene construct. Figure exemplifies the extraction and use of a coding ..., and have a clear vision of the final vector structure you want to achieve before starting your experiments...These overhangs are known as the common MoClo nomenclature. The overhangs are generated using type IIS ...

...solubility and stability (Figure 2).      Figure 2: Crystal structures of CRBNmidi alone (A) and...establishing CRBNmidi as a promising tool for future structure-guided drug design and studies of protein ...domains and is recommended for use in western blots (Figure 1). If you prefer to produce your own antibody ...chain are also available from Addgene!   Figure 1: Experimental validation for Anti-V5 [IPI-SV5...dimmer, showed greater photostability and also matured considerably faster, with expression levels comparable...comparable to GFP during early embryogenesis (Figure 3). mScarlet-I3 also offered clearer signals due to...autofluorescent tissues like the intestine. Figure 3: Expression of fluorophore-tagged GOLG-4 through...

...growing in requests faster than jGCaMP7 (Figure 5B).   Figure 5: A) GCaMP6, jGCaMP7, and jGCaMP8 viral...setup. Teams across Addgene have worked hard to ensure that the service has grown steadily, helped by ...our catalog had grown to 879 viral catalog items (Figure 1). The service caught on quickly in the scientific...distributed over 100,000 viral vector preps!   Figure 1: A) Cumulative AAV preps available in the Addgene... they’re delivering their payload to.   Figure 2: Numbers of AAV tools, by category, available...respectively, by substrates, light, or engineered ligands (Figure 2). Biosensors, which include calcium and voltage...explain why they are the most commonly requested (Figure 3). Over 400 biosensors were distributed in 2023...

...neuronal sub-cellular structures GFP is limited in its ability to label subcellular structures in the CNS. GFP...connections (Shepherd, 2004). Understanding the structure and function of these neural circuits is essential...experiments and visualize numerous molecules or structures at once (Shaner, 2007). In contrast to epitope...for the cells and affects the labeling of small structures like neurites. Like epitope tags, low FP expression...church of the Flying Spaghetti Monster; the smFP structure resembles images of the deity (see comparison ...processes. In contrast, smFPs can label fine neuronal structures with high fidelity and at lower concentrations...GFP. smFPs enable better labeling of these fine structures through their multiple, high-affinity binding...

...preparations are genome-containing (Figure 1).    Figure 1: Electron micrograph of an Addgene... by the temperature parameters. For example, we found that changing the annealing temperature of the PCR...viral vector solution, and generally these methods measure different characteristics of the solution. These... containing viral genomes. Physical titers are measured by quantifying the concentration of viral genomes... report the physical titer of a viral solution (Figure 2). Because physical titers are used for dosing...values mean and how they can be compared.   Figure 2: Label that comes on Addgene’s AAV aliquots. ...circled in red) report physical titer, which is measured by qPCR.   Physical titer is typically calculated...

... of temperature on total crop production and the production window. A two-way MANOVA could measure the..., you could test the impact of temperature, humidity, sunlight exposure time, and watering volume on total...power. Not sure if your data meets the assumptions of parametric tests? If you aren’t sure if your variance...outliers or data spread within those data sets. Measures of dispersion – standard deviation/error, variance...in a certain direction (i.e., you just want to measure if a sample is greater than a control), then a ...variable. A two-way ANOVA could test the effect of temperature and humidity on an outcome such as total crop... one-way ANOVA can test either the effect of temperature or humidity on crop production.   MANOVA The ...

... Figure 1: Schematic representations of the indicated unbiased genotyping options. Figure reproduced...other elements, including those relating to DNA structure (e.g. epigenetic modification, bulges) that may...conventional algorithms may not provide a full picture of the impact of CRISPR editing in the model cell...negatives Combining sequencing techniques can ensure validated experiments Unbiased detection methods...sequencing and in silico prediction should give a broad picture of all possible editing events in the genome; from...induced by CRISPR-Cas nucleases in human cells." Nature biotechnology 31.9 (2013): 822-826. PubMed PMID...reveals RNA-programmed Cas9 nuclease specificity." Nature biotechnology 31.9 (2013): 839-843. PubMed PMID...

...fluorescent protein mOrange2 (Figure 2). Figure 2: OCaMP structure and mechanism of action. Reproduced...brightness, ∆F/F0, or signal-to-noise ratio (Figure 3). Figure 3: One- and two-photon excitation and ... Figure 4: Calcium titration curves for OCaMP and various red and yellow GECIs measured with soluble...lasers available in your microscope setup. Figure 1: Excitation (dotted) and emission (solid) spectra...indicators spanning nearly the entire visible spectrum (Figure 1). The GCaMP series (e.g., jGCaMP8) remains the... yellow GECIs, OCaMP shows: Higher ΔF/F0 (the measure of signal change during calcium events), Greater...a CC-BY-NC-ND 4.0 International license. Figure 5: A) Simultaneous fluorescence (upper trace) and...

...downregulation. Figure 3 shows a histogram for a yes-no dot plot, like the one shown in Figure 1.  ...populations in the histogram plot in Figure 4 compared to the dot plot in Figure 2. Quantification using MFI makes...Before you read out your plots, you’ll want to make sure you’ve also adjusted voltage and compensated your...adjusting voltage: During data acquisition, make sure that the signal for the positive population is no...should overlap with the negative control population (Figure 1). Based on where the GFP- population ends, you... the total cells present in the gate.   Figure 1: Example of yes-no flow plot of the (negative...expression, low expression, and everything in between (Figure 2). These smears tend to occur when you are looking...

...Nishimasu H, Nureki O, Zhang F. Nature. 2015 Jan 29;517(7536):583-8. doi: 10.1038/nature14136. Epub 2014...Cas9’s crystal structure. “I thought maybe we could use information from the structure to guide and improve...technology. These libraries are lentiviral in nature, so please ensure that you are equipped and authorized to...genes, it is not possible to test each one and make sure it’s working. You can only do that screen with a... the SAM pooled library In a paper published in Nature in December, Konermann and her colleagues in Feng...-scale, knockout screening). As the Zhang lab's Nature paper reports, “Here we have shown that the SAM...Therapeutic uses in humans are also on the table. “In the future, there is potential there,” he says. "We have to...

...the purification procedure. Good indicators of DNA purity are the absorbance measurements at 230, 260, and...same. Before purifying a plasmid from a bacterial culture, it is important to consider your experiment.  ...Cloning grade DNA Transfection grade DNA Figure 1: Transformation introduces transformation grade...range). It can be extracted from small bacterial cultures (i.e. 2-3 ml) with or without commercially available...and DNA for transformation doesn’t need to be as pure as for other uses.   Cloning grade DNA Molecular...also dictate the required sizes of the bacterial cultures and commercial kits you will adopt for the extraction... cloning it is important that the preparation is pure from other nucleic acids (genomic DNA and RNA), ...

...applications.   Figure 1: Live-cell labeling strategies. The green structure represents a fluorophore... end up linked to whatever molecule you choose (Figure 1). Just express the fusion protein and add a specific...fluorophore or other useful ligand, gray structure is the protein of interest, and black is the fusion tag ...-tag construct can help you better detect dim structures or low levels of expression with the same imaging...super-resolution or single-molecule techniques.   Figure 2: Two-color super-resolution imaging of ER and... enables applications like multi-color imaging (Figure 2). If you’re working with purified proteins in...colors depending on the needs of your experiment (Figure 3). And using self-labeling tags makes updating...

...unique changes in their spectral properties upon exposure to a specific wavelength of light. PA-FPs can ...fluorescence, to a brighter fluorescent state by exposure to a specific wavelength of light. The switch ...switches from green to red fluorescence. mEosFP exposure to UV light causes an irreversible cleavage near...formation of mEosFP’s chromophore requires a temperature below 30°C and this is not ideal for experiments...emission of green fluorescence, but then further exposure to blue light also inactivates the chromophore...state. Dronpa can be reversibly reactivated by exposure to UV light. The fact that blue light both activates...photons are being emitted, making it easier to capture a clear image above background. The majority of...

... to ensure all plasmids pass our high-quality standards and contain the expected plasmid features.  ...based on SnapGene’s “Common Feature” library — with some additional features added by the QC team — and...the plasmid to ensure our assembly result is accurate The gene/insert or any major feature highlighted by...Nobody’s perfect! This is a platitude I’m sure we have all heard. It’s a truth that extends to both people...optimizing experiments, to false conclusions.  Figure 1. Potential points where DNA errors can be introduced... Figure 2. Addgene’s basic QC process. Created with BioRender.com. Once Addgene receives deposited...resolving a variety of QC issues. This process ensures that any potential sequence discrepancies can be...

... alert the day of the procedure. Mice move much more quickly than cell cultures do, and tired researchers... some nerves. Following safety procedures and practicing safety culture, however, can let you work with...risks and how to minimize them. Built-in safety features   When viruses were first being engineered as ...still enter (and thus infect) cells, this safety feature prevents the virus from replicating and spreading... including yourself as the researcher.   Figure 1: Separation of retroviral packaging components...precautions to take, and if something does happen, ensure you can receive appropriate treatment and/or monitoring...monitoring. Virus-specific concerns   In nature, some viruses are more dangerous than others, and the...

...present in the sample (APC in the case of Figure 1). In Figure 1A, the population needs to be shifted towards.... Let's look at an experimental example in Figure 2. Figure 2A shows the sample prior to compensation....the populations move, as in Figure 2, following compensation. Figure 3: Flow plot for a double positive... a compensation value between the two channels (Figure 1A). Applying compensation means you are telling...values later in your analysis, unlike voltages.  Figure 1A shows a single-color APC control, with APC and... a straight line and are only positive for APC (Figure 1B). For single-color samples, you can only change...would decrease the compensation value.   Figure 1: Compensation example with single color control...