... matches to your insert. Select the appropriate gene if it appears in the Entrez Gene list, as this will...would search for it in PubMed and choose the appropriate publication. This publication will be listed ...preprint. In the case of a preprint submission, we are happy to update the publication information once the manuscript...pass our standard QC process and the MTA has been approved. The spreadsheet option is recommended for ...contain epitope tags or fusion proteins that are appended to new inserts through cloning. *Note — If you...complete these pages sequentially by entering the appropriate data and pressing "Save and Continue to Next ...around to the different pages by clicking on the appropriate boxes at the top of the page. Pro Tip! Be sure...

...the filament from (a-b), showing the appearance and disappearance of 649-mDia1 or 549-CP (upper, middle...block its catalytic activity and instead become trapped in a covalent bond with its substrate. Then, all...having to clone a whole new construct for every application. Plus, the photophysical properties of chemical...advantages over fluorescent proteins, opening up new applications.   Figure 1: Live-cell labeling strategies...acids 1 Various Many Complicated   Applications Organic dyes are typically both brighter and...self-labeling tag and a fluorescent protein) enables applications like multi-color imaging (Figure 2). If you’...Actin, cyan), with formin (649-mDia1, magenta) and capping protein (549-CP, yellow) at the tip. 649-mDia1 ...

...depending on the conference you’re attending and who approaches you. You might also have cross-team projects ...communication. Although these real world examples did not happen in the lab, they have parallels to collaborative...types or purposes of each given container, when applicable Any other relevant information Prescribing an...misinterpretation, that misinterpretation was definitely happening. Our solution for this problem was the creation...lab setting, where student and postdoc turnover happens regularly and it behooves everyone in the lab to...potentially ambiguous terms. Defining words and applying annotation can be particularly useful in academic.... But if you implement it well, here’s what can happen: More effective meetings. When everyone at a meeting...

...truth is, there is a lot of software development happening at Addgene under the hood. A significant portion...this can lead to wildly different ideas of how to approach a problem, but it is these very differences that...similar cognitive biases and this would become apparent in the software we develop. We would all have ...framework is based on an iterative and collaborative approach to development. Scrum teams plan and break up ...are also able to consider different technical approaches that may come up during discussion.  One example...perspective which in turn helps us decide how to best approach it in our code.  Sometimes while thinking through...that the clarifications lead to the story being scrapped or broken up into smaller stories. Since this ...

...virus If having happy cells is the first step of a successful lentiviral infection, happy virus is the second... express a gene of interest. While the general approach for using lentivirus, infect and select, seems... cell lines, ELISA, fluorometry, or PCR-based approaches to detect even low levels of contamination. For...Uphoff et. al. outline a do it yourself PCR-based approach complete with freely available positive controls...old. Ensuring that you are working with healthy, happy cells is one of the easiest ways to improve both...envelope. One drawback of the multiple harvest approach is that it tends to result in a higher volume ... freshly collected lentivirus for downstream applications. While this ensures that the highest titer virus...

...non-refundable fee is applied to cover processing costs. Once the application is approved, Addgene then ships...excited to announce we now offer an export license application for rabies materials! Some of the materials in...international requestors. We also offer export license applications for VSV and SARS-COV-1 plasmids. Export control...license process    An export control license, once approved, allows Addgene to ship plasmids or viral vectors...will need to provide a brief description of the applicable research project when purchasing export-controlled...export-controlled materials. Addgene will then apply for the license for distribution to the requester’s organization...offered VSV and SARS-COV-1 plasmids under this application process since May 2020. We find this process ...

...led to their application for basic and translational research.  Basic research applications of scFvs: ...blood clearance, which is useful for imaging applications; better tissue penetration which is useful for... for therapeutic and imaging applications; and reduced immunogenicity when administered in vivo due to...from blood can be a drawback for therapeutic applications where longer retention times often increase ...laborious.  To avoid this, one or more of these approaches are often used for scFv expression in bacteria...since it’s genetically encoded.   Translational applications of scFvs: Just like antibodies, there are many...2019).Many of the scFvs designed for oncology applications are reformatted as bispecific molecules which...

...fits all When choosing an appropriate antibody first look at the application-specific data provided on...your specific application. Consider also that there may be variations within an application. Take immunohistochemistry... a linear antigen tend to work very well for applications that denature a protein to its primary (linear...while the website highlights a variation of your application, a publication successfully used it in a similar...plan to. To secondary or not to secondary?  The application will also determine whether you should use an... as it will help to amplify the signal. Some applications use either direct (primary-conjugated) or indirect... are secondary considerations to validation, application specificity, and experimental considerations ...

...your experiment (all well validated for your application, of course!) When the stars are aligned and the...factors that affect isotype choice. Target and application should be considered The first question you may... a strong signal and to work in a variety of applications. The desired specificity of your secondary antibody...affect my experiment?” Well, it depends on the application and target. If you are running a basic western...target is expressed at very low levels or your application requires a high degree of affinity, then species...considered when choosing an antibody. In some applications probing species-on-species, i.e. using a mouse...easily distinguishable from each other. In a direct approach, that is simple - but what about when you need...

...many applications, it is the only comprehensive method available. Targeted sequencing approaches More...differently. For a comprehensive approach, RNA-seq would be an appropriate method for an activator or repressor...lower them even further. While not all of these approaches will work for every experiment, at least one ...where only localization of the Cas occurs. 2 gRNA approach Off-targets that come from mutations arise from...More targeted alternative sequencing approaches have been developed. Cas ChIP-Seq, where only DNA sites ...especially when DNA mismatches are involved. Two other approaches have been used - GUIDE-Seq and integrase-defective...site-specific detection downstream. The con to these approaches is that NHEJ must perform the repair to allow...

... to be placed immediately upon request approval. Once approved, the request will remain in your account... in this blog post.     When will the expansion happen?  You will be able to request Packaged on Request...request will be reviewed by Addgene for legal approvals and technical compatibility with our production...process will take 1–2 days, and we anticipate approving the vast majority of requests.  If the vector ... request process.  Ordering If your request is approved, you will receive an email notification that your...is ready to order. An “Add to Cart” button will appear with the request in your Addgene account.  Orders...viral preps will be available at launch.  What happens after I order?  Once an order is placed, the Addgene...

...distinction of neighboring cells in imaging applications and the mapping of neuronal projections to their associated...while minimizing confounding secondary effects? An approach described as “knocksideways” (a British idiom ...domains, such as FKBP and FRB. The cell does not appear to mind extraneous proteins bound to its outer ...protein from expression to degradation. Chemical approaches may require expensive peptides or small molecules...and Technology Austria, decided to take a novel approach to control for these effects, drawing on the rapidly...optogenetics. Using a rational protein engineering approach, they designed Opto-RTKs, which activate signaling... on exposure to low-intensity blue light. This approach relies on the incorporation of light-oxygen-voltage...

... can happen at the national level within the executive branch or in Congress, or can also happen at the... there will be posts about virtual meet ups and happy hours that you can attend to help build relationships...arriving here in DC, is that so much policy actually happens outside of the government itself. You can work ...The “little p” policy is non-governmental work happening at large organizations who have to develop policies...who decides how they are going pursue regulatory approval for a new product or a well-endowed philanthropy...Program to learn more about where science policy happens.  When do you do science policy? Many science ...Project for some example proposals and consider applying for their accelerator for some hands-on guidance...

...sense and appreciate this – and they will be happy too. On the flip side, if you are not happy at work,... If you are happy at work, you will find balance. If you enjoy working and come home happy in the evening...actions we can take to help ourselves can be an appropriate first step. Check out Joanne's Reddit AMA Managing...turns going; that way you can have someone there clapping more often than not. You want your family and ...tools and tactics to plan your work and make this happen. Be realistic and: Your co-workers will learn ... about their safety when they leave home which happens very, very fast! I will never forget the time my...figure out a way to keep being ourselves and to be happy. Building your plan to live a good life may be the...

... are approximately 4.1 kb in length, allowing them to be packaged in AAV.   Split AAV approaches  While...germline genome editing, but in vivo gene editing approaches have been limited by problems with Cas9 delivery...packaging capacity of AAV (~4.7 kb). While this approach has been proven feasible, it leaves little room...potent Cas9 protein, the Zhang lab took a different approach. They analyzed over 600 Cas9 orthologs and found...be divided into two groups: longer orthologs approximately 1350 amino acids in size, which includes SpCas9...SpCas9, and shortern orthologs approximately 1000 amino acids in size. From the pool of shorter orthologs...effects of SpCas9 and SaCas9, Zhang’s group used an approach called BLESS (direct in situ breaks labeling, ...

...platform offers a promising approach for experimental therapies and medical applications.  Find DIRECTED plasmids...Figure 1: Choosing the best PE6 variant for your application. Image reused form Doman and Pandey et al., 2023...REcipient Cells Through Envelope Design). This modular approach combines an interchangeable set of fusogenic and... CreVLPs, and eVLPs for in vivo and in vitro applications.   Figure 2: Modular features of DIRECTED...interactions or map a local proteome, but some applications have been challenging or inaccessible. The Ting...  Qin, W., Cheah, J.S., et al. (2023). Dynamic mapping of proteome trafficking within and between living... from different promoters, specific time points appear as fluorescent bands in the fiber, resembling a...

...fluorescence can be visualised. Another approach was to apply the same concept to lateral flow strips ... Suresh from Iowa State University. Since its appearance, SARS-CoV-2 has spread to almost every part of... required: 37°C heat block and 62°C heat block Approach: Viral RNA is extracted and subject to isothermal... required: 37°C water bath and 42°C water bath Approach: RNA is extracted from a sample and subject to...cleaved. Cleavage results in fluorescence and this approach can be incorporated into a lateral-flow readout...blue light illuminator or UV light illuminator. Approach: Highly similar to the SHERLOCK diagnostic test...microliter Time: Not mentioned in the paper, likely approximately 1-2 hours Equipment needed: Mini PCR machines...

...is an application in which antibodies that have been joined to fluorescent molecules are applied to a ...immunofluorescence There are a couple different approaches to be familiar with when talking about IF, direct...BioRender.com   Indirect IF is a much more common approach than direct IF, so we will focus on this technique... Primary Antibody This is the first antibody applied to your sample, which will bind to your target ...Unsurprisingly, this is the second antibody you apply, which binds to the first antibody and is conjugated...distribute antibodies validate them in a variety of applications, including IF (or IHC or ICC), and will share... time to research antibodies, protocols, and appropriate controls and you will be well on your way to ...

...sample of your protein of interest (POI). This approach is reliable, but it’s overkill in many cases. ...than “condition 2.” By eye, the loading control appears about the same between the two conditions. But ...detected with an antibodies, just like the POI. This approach is so common that we’ve used it for our cartoon...cells. This means that at sample concentrations appropriate to detect a less-abundant POI, the loading control... control aren’t too far apart, they may have overlapping linear detection ranges, allowing you to accurately..., 2008; Gilda & Gomes, 2013). Depending on the approach you choose, your loading control measurement will...on the same gel or membrane (depending on your approach) as your final blot. Quantifying the entire lane...

...Blocked by overlapping methylation Not affected BsaI Not affected Blocked by overlapping methylation...Blocked by overlapping methylation HpaI Not affected Not affected Blocked by overlapping methylation...Blocked by overlapping methylation Not affected Not affected MscI Not affected Blocked by overlapping methylation...Not affected Blocked by overlapping methylation XbaI Blocked by overlapping methylation Not affected...methylation Not affected ClaI Blocked by overlapping methylation Not affected Not affected DraI Not affected...