We narrowed to 820 results for: APP

...knockout approaches. In these cases, we would recommend a knockdown approach instead. In this approach, you...using one or more of the approaches outlined below. Knockout cell line approach Genetic knockout-based...Knockdown approach For cell types where knockout studies are not feasible, an alternative approach is RNA... the target of interest. This approach maps onto the orthogonal approach within the five pillars and is...knockout/ or knockdown approach should be preferred. The overexpression approach is therefore best suited... use of orthogonal approaches, cell treatments, and independent antibody approaches in different assay...target-of-interest in your assay and sample-of-interest. The approaches available broadly map onto the five pillars of...

...assume that their advisor alone applies for these competitive grant applications; however, in some cases, your...the grant application process. When my PhD advisor first asked me to write a grant application due the ...mind that universities require application submission in-house approximately 2 weeks before the RFA due date...the next round of applications. Step 5: Keep writing! Each written grant application builds upon the ... project grant application" by the NIH Still want more information on the application process? What types...writing, volunteered to take the lead on various applications throughout my graduate career, and helped receive...opportunities to convince others why a project is the best approach to a given problem, Empowers the writer, And...

...intended application - Remember, just because your antibody performs as expected in one application does ...Strategies This approach compares target expression of antibody-independent approaches to your antibody-dependent...Genetic Strategies This approach uses a gene knock-down or knock-out approach to show that an antibody...protein. This approach is ideal for validating antibodies for immunocapture applications and is amenable...pause and make sure that your antibody has been appropriately validated so that you can have confidence in...in your results. But what is appropriately validated? This post will go over some of the general considerations...strategies for antibody validation in different applications. Here, I want to highlight the general suggestions...

...allow you to approach that problem in a novel way. Finally, engineering new applications to solve problems...good quality application. Even following this advice, only about 50% of my grant applications have been ...do need to be able to research and write your application, then give it to someone else to read and / or...it). It’s easy for reviewers to spot when an application is rushed (whether the idea isn’t properly researched...researched / developed or the application is full of typos) and these are unlikely to get past the initial...someone else in the department) to read over your application and give you feedback. They’ve most likely been...alterations before submitting - handing them your application 3 days before the deadline isn’t going to be ...

...of four split AAV vector approaches. 1. Overlapping  Overview: The overlapping strategy has a region of...directed repairs (HDR). Split AAV vector approaches Split vector approaches increase the size of the gene delivered...  3. Hybrid  Overview: The hybrid approach combines the overlapping and trans-splicing methods. The AAV...has a slight advantage over the overlapping and trans-splicing approaches since there are two ways for a... genomes is the basis of several split vector approaches developed in the early 2000’s which allow researchers...is not yet known. Drawbacks:  Generally an overlapping split vector has equal or higher transgene expression... on the rate of transgene expression with an overlapping AAV since both of these variables influence whether...

...is this application - the creation of sequence-specific knockins - for which the AAV approach is most ...for a given experimental approach. First, unlike CRISPR, the AAV-based approach is only able to target ...simultaneously in the same cell, are more appropriate for a CRISPR-based approach. Second, the AAV preps generally...limitations, the AAV approach is best suited for tissue culture based approaches that require modification...researchers could apply this technique to efficiently deliver large repair templates for applications that require...working to develop high efficiency gene targeting approaches improved on this finding with the addition of...challenging to develop high efficiency CRISPR-based approaches for the generation of knockins via homologous...

.... This is the direct approach to immunoimaging assays. But if the direct approach isn’t giving you enough...   Figure 1: The direct approach (left) versus the indirect approach (right). Primary antibodies ...varies across applications. Western blots, for instance, almost always use the indirect approach, while flow...immunocytochemistry may all use either the direct approach or indirect approach, depending on the requirements of the...interest, it may be time to look at the indirect approach - which means it’s time to talk about secondary...binds to the primary antibody. In the indirect approach, the signaling molecule is conjugated to the secondary...antibodies can bind to a single primary antibody, this approach can greatly increase the sensitivity of an assay...

..." are summarized below. Check Out Erik Snapp's Full "Applying for a Faculty Position" eBook Here 1. Have...  Erik Snapp just released version 3.4 of his eBook, How to Get a Faculty Position. This edition has ...along with new thoughts on research statements. Snapp, who is always looking for ways to improve his eBook...suggestions, please do not hesitate to email me at snappe@janelia.hhmi.org." You can find his original blog...years ago, I decided to write a "how to" manual on applying for faculty positions in biomedical science. My...talents and ideas. Some key points from my ebook, "Applying for a Faculty Position: the View from Both Sides...experts in your field. Not true. Even if you are applying to a department in your specialty area, it's a...

...safe for clinical applications. Excisable Cre-lox lentiviral vectors: This approach is the same as using...of excision, this approach is still not considered safe for therapeutic applications. piggyBac (PB) transposon...This is considered a safer approach for developing therapeutic applications, but it can take a long time...time, scientists have used almost all available approaches for generating iPSCs. The generation of iPSCs...Retroviruses are not considered safe for clinical applications.     Lentiviral: Lentiviral vectors infect dividing...dilution via cell division. A drawback to this approach is it has lower levels of efficiency at generating...express reprogramming factors for therapeutic applications. Sendai viral vectors: Sendai virus is a single...

...relationships. However, most job seekers will and should apply for jobs posted online as one part of their job ...Utilizing existing relationships as part of the job application process can dramatically increase the chances... of a resume can be crucial for a successful application. It takes time to prepare an excellent resume... Many job postings will result in hundreds of applicants the day they are posted.  If your resume is not... here for more on timely submission of a job application. 3) Read job descriptions carefully Be open minded...have to have all of the listed qualifications to apply for a position.  A job description is usually a ...wish list and the hiring manager knows that no applicant is likely to meet all the criteria. Note the criteria...

...skipping approach in mice, DMD researchers are very excited. This approach may also be applicable to a number...success in basic research, researchers are eager to apply it in a clinical setting. CRISPR is often used for...), an X-linked recessive disorder affecting approximately 1 in 5000 males. DMD is caused primarily by ...very large (79 exons), but much of the sequence appears to be nonessential. Within the mutational hotspot...so researchers have set their sights on another approach. Since shorter forms of dystrophin can still be...researchers have begun to explore genome editing approaches for exon skipping. Addgene depositor Charles ... removal rate of exon 51, which resulted in appropriately localized dystrophin. In a subsequent study,...

...genes provided by the user. This approach is useful for several applications as will be discussed below. The... screen approach is to use smaller libraries targeting subsets of genes. With such an approach, you lose...custom CRISPR screen and the appropriate phenotypic readout, similar mapping could be rapidly performed ...challenges with a custom screen approach is to identify an appropriate list of genes to target. Thus, ... involved in a studied cellular behavior. This approach is particularly suitable for in vivo screens, ...drugs. Known drug targets  Another interesting approach is to design screens against known druggable targets...preferentially those for which validated inhibitors or even approved drugs already exist. If you are lucky and find...

... tools for familiar applications. But how does one invent a tool for applications that don’t even exist...and develop potential applications.  Andrew is a frequent collaborator and is happy to give advice and support...phenomenon, improve the tool, and develop new applications for magnetoresponsive proteins. Motivated by...organisms) are opaque, which makes most biomedical applications of these tools difficult or impossible. But ...tool with magnets would enable new research and applications that don’t yet exist, but it wasn’t clear that...molecules like proteins with magnetic fields generally appears to be weak — if they exist at all. Andrew and Maria...fluorescence intensity decreased whenever the magnet approached the sample (by less than 1%) and recovered after...

...single model. The first approach can lead to new directions, while the second approach could terminate with... Many thanks to our guest blogger, Erik Snapp! Erik Lee Snapp received his PhD in Molecular Microbiology...This post was contributed by Erik Snapp, the Director of Student and Postdoctoral Programs at the Janelia.... Given this large sum, one might expect that applicants would work for months to hone a chalk talk. Yet...preparing for both types. Your research vision is your approach to resolving a significant knowledge gap in your...has unclear utility. You cannot predict what will happen. Without the prediction, you have no hook. A battery... you need to be able to describe experimental approaches in terms of the question, the methods, the tools...

...our guest bloggers Erik L. Snapp and Lindsey M. Costantini! Erik Lee Snapp received his PhD in Molecular... post was contributed by guest bloggers Erik L. Snapp and Lindsey M. Costantini. "You underestimate the... of a mystical "Force," this quote is equally applicable to many microscopy experiments with fluorescent...not fluoresce. A dark population is not readily apparent and can confound quantitative imaging experiments...for use in cytoplasmic environments. However, approximately 40% of human (and most eukaryotic) proteins ...have reported that FP cysteine residues form inappropriate disulfide bonds in the secretory pathway (Figure...localization. For example, the formation of inappropriate inter-chain disulfide bonds between FPs blocks...

...developed and applied. The current pandemic serves as a striking example of the applicability and importance...research. I applied, hoping that the experience would improve my scientific and application writing skills...science (as opposed to translational, biomedical, or applied sciences), people conduct research to answer questions...revelation. Science in any field can generally be applied to improve our lives and solve global problems....grants Varying degrees of engagement with what happens in the lab Official title is typically Professor...the people around you. Be proactive Rather than applying only to posted job offerings, the best research...ties into the previous one too. I would recommend applying for multiple positions—don’t put all your eggs...

...received 116,224 employment based green card applications and approved 91,023 during the first 3 quarters of ...options discussed in this post benefit from the applicant thinking about the process early, keeping good...is a numerical quota of 85,000 total H-1B visas approved each fiscal year: 65,000 visas plus 20,000 reserved...occupation" as requiring theoretical and practical application of a body of highly specialized knowledge in ... April 1 deadline for companies to submit an application on their behalf.  If an individual obtains a ... cap subject H-1B (last year less than 30% of applicants received sponsorship) then he or she can start...1B petitioners are exempt from the cap if the applicant works at institutions of higher education, a related...

...real-time PCR (4). Restriction mapping and modern sequencing Restriction mapping informs on DNA sequence through...useful applications and have informed many current techniques. For example, restriction mapping led to...historical techniques: southern blotting, restriction mapping, and sequencing gels, as well as their modern equivalents...comparison to a control sample. Southern blotting applications range from confirming the results of cloning...gene that causes sickle cell anemia. The mutation happens to destroy a sequence recognized by the restriction... was commonplace, researchers used restriction mapping to investigate DNA sequence content by comparing...of the DNA. Starting in the 1970s, restriction mapping was commonly used to map DNA and examine genomes...

...zebrafish embryo has been mapped using single cell RNA sequencing approaches (Harland, 2018). Several ...investigations. Genetic resources for zebrafish A common approach to unravel the gene networks that drive biology...DNA into the genome is useful for a variety of applications. For example, specific tissues such as blood... regions is also possible using an alternative approach that exploits the bacteriophage integrase phiC31...useful, I’ll outline two recent lineage tracing approaches that led to important insights in developmental...tracked using microscopy and advanced computational approaches (Keller et al., 2008). This microscopy-based ...pathways control this process. Additional zebrafish applications and conclusions Zebrafish are also popular outside...

...subsequent, repeated activation), this is an approximation of reward or the sense of pleasure that the ...EthoVision. Consult the literature to find all the applications of optogenetics relevant to your field. Ultimately...behavior, certain technical considerations still apply. Testing stimulation First, In lieu of electrophysiology...stimulate them for 30min to an hour and observe what happens. You never know how stimulation of a particular...optimization, turn on the laser and watch what happens. Who knows, you may observe something that could... discovery. Monitoring behavioral output The appropriate behavioral readout for your optogenetics experiment...brain region that may be important in memory, an appropriate behavioral test might be contextual fear conditioning...

... a bioinformatics approach examining genomes possessing CRISPR systems with apparent self-targeting. Two... plasmid or group of plasmids' functions and applications. We hope that these articles make it easier ...includes four promoters, six fluorophores with nonoverlapping emission spectra (N- and C-terminal tags for...Targeting and manipulating interneurons using a viral approach Genetically modified organisms like transgenic...successfully targeted via viral particles, an effective approach to target specific neuronal subtypes was not available..., Gordon Fishell’s lab has developed a new AAV approach using regulatory elements that restrict expression...tools in virtually any vertebrate. This novel AAV approach allows you to target and functionally manipulate...

... 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...

... use one of two common approaches — columns or magnetic beads. Both approaches involve binding the plasmid... plasmid preps are the same. Your downstream application will dictate how much DNA you need to recover...recover and the level of quality required. Some applications require very little DNA and you can get away with...with slightly lower quality preps, while other applications require large amounts of high quality DNA. Figure... a High Quantity/Low Quality mindset for any applications. DNA quantity is fairly self explanatory and...possibility of damaging the plasmid DNA itself, which can happen if you leave the sample in a lysis buffer for too... the DNA from the substrate. In a column-based approach, the substrate is a silica matrix fixed in a column...

...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 ...

...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...

...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...

... allows components to be swapped out, potentially broadening its applications. Find ATLAS plasmids here...Addgene’s own AAV6 and AAV11 packaging plasmids Mapping anterograde synaptic connections with ATLAS Bright...between mGold2s and mGold2t depending on the application. Figure 1: Super-resolution imaging...proof-of-concept, they believe that the strategy will be applicable with other TnpBs and in other plant species, ...available for tracking cell lineage, different applications require careful selection or design to ensure...Alejo Rodriguez-Fraticelli recently developed an approach called Simultaneous Tracking of Recombinase Activation...BY license. Libraries are available with T-Sapphire, EGFP, or mScarlet reporters, so you can choose...

... 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...