...users to search for protocols in protocols.io, and then link to a selected protocol and/or import and convert...powerful example is the integration with protocols.io, the protocol sharing repository. As illustrated in... possible to re-export a protocol which has been edited back to protocols.io. Thus, RSpace becomes part...management software, and method repositories like protocols.io. These tools are essential to the workflows ...enhances the sharing and reproducibility ecosystem protocols.io makes possible. Connectivity to open source ...scientific sharing Resources on Addgene.org Find protocols of molecular biology techniques Find plasmids ...

...overcome these woes with photoactivatable and photocleavable gRNAs. Photocleavable guides are generated by...light. The photoactivatable counterpart to this system utilizes a gRNA which is photocaged and unable ... by introduction of a single photocleavable 2-nitrobenzyl linker in the 20-bp targeting region of the ...enzyme to pair with your guide and our available protocols to get your CRISPR experiment moving. Happy editing..., Y., Wu, B., et al. Cas9 deactivation with photocleavable gRNAs. Molecular Cell 81, 7, 1553-1565 (2021..., E., Satyapertiwi, D., Ramel, MC., et al. Photoswitchable gRNAs for Spatiotemporally Controlled CRSIPR-Cas-based...

...with, the protocol will slightly differ. For example, Girardia sp. and Dugesia dorotocephalacan can be fed...available planarians: Girardia sp., Dugesia dorotocephala, Phagocata morgani, and Phagocata gracillis....from the Sánchez lab describes the plasmids and protocols required for both WISH and RNAi experiments, as...class” which provides more detailed materials and protocol resources. Find Sánchez lab Plasmids Visualizing...tissue. Once the riboprobes are created, the WISH protocol involves preparation of the tissue sample, several... the translation machinery. In the Sánchez lab protocol, dsRNAs are delivered by feeding them to planarians...riboprobes and dsRNAs into PR-T4P. The Sánchez lab protocol also provides step-by-step instructions for the...

...emits green, yellow and orange photons while PE emits yellow and orange photons. If FITC and PE were being...or following labor-intensive limiting dilution protocols? If this sounds like you, then flow cytometry ...captured by detectors. The strength of the emitted photons varies from cell to cell depending on the level...fluorochrome-stained protein will emit a stronger photon than a cell expressing low levels of the protein...then follow the typical intracellular staining protocol. Sorting cells with FACS In addition to the fluidics... antibody. When following an indirect staining protocol, you should also include a secondary antibody ...primary antibody has been omitted from the staining protocol. This control allows you to detect background ...

...scientist and photographer based at the University of Queensland, whose outstanding marine photography has attracted...” Daniel went on to discuss his work: “Macro photography has been my passion for a while - a tool to expand...life cycle of the creature they deal with.” So photography can be used as a way to better understand and...scientist and non-scientist alike. “The other thing photography has shown to me - it requires an awful amount...data, publish in time, defend the thesis, etc. Photography is critically important in many fields of biology... can be found on his website http://www.microworldsphotography.com/. Another artist who spent his life...artists can compete in the Nikon Small World Photomicrography Competition - a 40 year old international ...

... accurately run a protocol.   Figure 1: Evolution of a Western blot protocol, from a full page ...checklist for everything you do in the form of a protocol, and you can add in personalized steps like “check...re-sterilize now.” Have a physical copy of the protocol when you work and make a checkmark next to each... it anyway.  You may find that after running a protocol a few times, you no longer need the full checklist...check things off every 3 steps, or you rewrite the protocol and consolidate several steps into one. Soon you...as you run your experiments. Try drawing out a protocol as you read it, or putting a diagram up on your...may seem counterintuitive to add more time to a protocol if you’re in a hurry, but let’s consider it this...

...engineering channelrhodopsins to have improved photocurrent properties: The sequence and structures that...channelrhodopsin, as well as predict the channelrhodopsin’s photocurrent strength, wavelength sensitivity, and off-kinetics...predicted channelrhodopsin sequences would have photocurrent properties best suited for less-invasive optogenetics...All thirty were functional and their measured photocurrent amplitude, wavelength sensitivity, and off-kinetics...neurons expressing ChRgers produced stronger photocurrents at low light intensity than existing channelrhodopsins...response. This result showed that ChRger2 stronger photocurrent properties make it better suited for systemic...non-invasive methods to the neurons in the right secondary motor cortex (M2). Since M2 is located on the surface ...

...existed! Addgene recently started automating a few protocols to make standard lab processes easier and more...own lab. Automating bacterial transformation One protocol that has been automated is high throughput transformation... machine that would improve our transformation protocol and, after looking at a few options, we decided... to use the FXP to automate and optimize more protocols in the future - something that will be key for...same amount of time by hand! Setting up a basic protocol on the Biomek FxP is pretty simple. By clicking...perfectly. Working with Qiagen, we developed a protocol to run up to 192 samples at a time. It follows...contamination between samples. With this automated protocol, we can prep two 96 well plates in just under ...

...It was a bit intimidating, but I had my lab’s protocols and some great co-workers to help me out. Even...writing AAV production tricks into the margins of my protocol. While these tips weren’t critical to the experiment...HEK293 cells. Check out the Addgene AAV Production protocol for more details. Pro tips I’m not going to lie...enough AAV. Compared to the Addgene AAV Production protocol, I take a lazier approach for harvesting my AAV...the AAV Iodixanol Gradient Ultracentrifugation Protocol for more details. Pro tips Watch this AAV purification...analysis. Refer to this Addgene AAV Titration by qPCR Protocol for more details. Pro Tips When calculating ...vectors for in vitro and in vivo use. Current protocols in molecular biology, Chapter 16, Unit 16.25. ...

...focus light. Therefore, there is much less photo-bleaching or photo-damage than in conventional laser scanning...order to improve axial resolution and reduce phototoxicity for imaging live specimens, they combined light... through the specimen, which often leads to photobleaching.  Furthermore, point-by-point scanning along...light scattering and absorption.  Finally, the photomultipliers used to detect light in confocal microscopes... Bradke, M. Sheng, H.-U. Dodt. (2012). Nature Protocols, 7(11), 1983–1995. Pubmed. K. Chung, J. Wallace...Tomer, L. Ye, B. Hsueh, K. Deisseroth. (2014) Nat Protoc., 9, 1682-1697. Pubmed. C. Soderblom, D.-H. Lee...

...categories of fluorescent proteins Photoactivatable / Photoconvertible: These proteins can switch their ... fluorescence. Examples for such photoactivatable or photoconvertible proteins are PA-GFP, Dendra2, and...0.1) and this should be taken into account. Photostability can be affected by experimental parameters ...excitation light intensity, pH or temperature). Photostability: Fluorescent molecules gets bleached (i.e. ...after prolonged exposure to excitation light. Photostability can be as short as 100ms (EBFP) or as long ...most FPs it is a few seconds to a few minutes. Photostability can be affected by experimental parameters ...

...based on the location of the ‘A’ relative to the protospacer adjacent motif (PAM). ABE7.10 can edit target...'A' if it lies in a small genomic window: at protospacer position 4-8 counting the ‘NGG’ PAM as 21-23 ...closer to the PAM, i.e., positioned at 8-10 in protospacer, ABE6.3 or ABE7.9 are better suited (Gaudelli...two Bsa1 sites to clone the single guide RNA (protospacer+scaffold) in a pol-III (OsU3) promoter driven...tRNA-gRNA (PTG) gene system to clone multiple protospacers which enables us to target more than one genomic...smaller vectors suitable for transient assays using protoplast transformation. Cytosine base editors for plant...placed in 2-6 bp and 4-8 bp, respectively, of the protospacer. And if the target cytosine is closer to the ...

...binding. Photobiotinylation Photobiotin can be used to non-selectively label DNA and RNA. Photobiotin acetate...biology for an array of different techniques and protocols. In this post we will discuss the natural role...consists of a biotin molecule attached to a photoreactive arylazide group. Under strong visible light,...nearby nucleic acids (McInnes et al., 1990). Photobiotinylation is sometimes easier and less expensive than...C., & Symons, R. H. “Preparation and uses of photobiotin.” Methods in Enzymology 184 (1990): 588–600. ...biotin ligase and monovalent streptavidin.” Nature Protocols 3.3 (2008): 534–545. PubMed PMID:18323822. PubMed...

...other antibodies. But, once you do find the right protocol, multiplex IF can give you impactful and beautiful...manufacturer’s recommendations.  As you optimize your protocol, always remember to include controls to show that...experiment will have a significant impact on the protocol you use and the ultimate outcome of your assay...share some of their results and corresponding protocols. Other researchers may also have tested an antibody...website. It is important to review the data and protocols available for each antibody that you consider ...how reliable the antibody seems and what sort of protocol you will need to use. (To learn more about antibody...home?). Take your time to research antibodies, protocols, and appropriate controls and you will be well...

...increased brightness, sensitivity, and photostability under two-photon illumination than previous GEVIs. Some...called JEDI-2P (Liu et al. 2022) optimized for two-photon imaging. They use a unique and powerful high-throughput...based on performance metrics specifically for two-photon microscopy. The resulting JEDI-2P has faster response...increased voltage response, brightness, and photostability. It enables sustained deep-tissue imaging in...recording using a fast indicator evolved for two-photon microscopy. Cell. 2022 Aug 16. pii: S0092-8674(...iGECInano displays better response kinetics and photochemical characteristics than previous NIR calcium reporters...candidate biosensor variants for performance under two-photon fluorescence lifetime imaging. B) LiLac responses...

...signal-to-noise ratio Less chance of phototoxicity Reduced photobleaching Simplified, long-term single-molecule...enough signal. This workaround risks cellular phototoxicity and eliminates the possibility of long-term ...a higher signal-to-noise ratio with a lower photobleaching rate. Given the power of SunTag, they attempted...- they even managed to track run lengths of the motor protein kinesin across microtubules. Tanenbaum ...

...-fold more photostable and as bright to mVenus, the parent YFP, and is the most photostable YFP to date...improved activity as compared to other recombinase/photodimer pairs. Created with BioRender.com In their... publication, Mary Dunlop’s lab generated a photosensitive Cre, Opto-Cre-Vvd, by splitting and joining...joining the protein regions with Vivid (Vvd) photodimers. Upon the application of blue light, Vvd dimerizes....ncbi.nlm.nih.gov/32675376/ mGold, the most photostable yellow fluorescent protein to date By Justin ...contribute to experiments that require high photostability, such as time-lapse imaging or with high light...

...well-established protocols for their production and purification that yield high titers (see also AAV protocols for...neuronal connectivity. These methods are less neurotoxic and better compatible with other neuroscience...system where it replicates, spreads, causes neurotoxicity, and is lethal if left untreated (for review...AAV), have been used for tracing and visualizing motor, visual, and auditory neuronal circuitry in vivo... project but neurons were traced with AAV2. Two-Photon imaging was used to generate a 3D finemap of neurons...WGA-Alexa Transsynaptic Labeling in the Phrenic Motor System of Adult Rats: Intrapleural Injection versus....M., and Mooney, R. (2017). Identification of a motor to auditory pathway important for vocal learning...

...for a western blot protocol? Check out Addgene's western blot protocol and protocol video!  Antibodies...Addgene.org Addgene's Western Blot Protocol Addgene's Western Blot Protocol Video References Aldridge, G....overwhelming that you might just stick with the protocol your labmate handed you — after all, it worked...you'll be able to understand how to modify the protocol to one that works for you. There's a lot to cover...However, they also add time to an already lengthy protocol, require the use of hazardous chemicals, and are...consistent. Precast gels, while expensive, reduce protocol time, can be stored at 4 °C, and are very consistent...

...sections: Protocols To complement our written protocol materials, we have also created video protocols.  These...navigate various lab processes.  Our most recent protocol video detailing Streaking Plates for Single Bacterial...Check Our Our Protocol Pages Browse our Plasmid Reference Pages ... protocol video...