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TypeBlog Post...You need the support of your institute to do this one. Universities will usually only support tenure track...Further Evidence (RFE). Letters of support are the only opportunity that you are given to present your...and so on. Candidates should look broadly at opportunities to develop skills related to their areas of ...must have both substantial merit and national importance; (2) The foreign national must be well positioned...evaluating an NIW, USCIS will also focus on the importance of the individual’s work in their field and the...working under in the lab or has won, awards, media reports, and membership in groups related to the foreign...This criterion is best proven through letters of support from experts in the field. You must also be able...
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7 Tips to Secure a STEAM Internship This Summer
TypeBlog Post...STEAMid. Internships open doors to a web of opportunities that the classroom cannot offer. Furthermore... not deter any student from applying to the opportunities of their dream. To help students focus on the...below: 1. Eligibility check Scrutinize every opportunity prior to starting an application to assess your...unless you are applying for a Federal Government opportunity, which tends to be more detailed. Beyond the ...draft a personal statement, which is the perfect opportunity to paint a comprehensive picture of your academic...statement will pay dividends. This is the only opportunity where you get to impress the judges on your ability...your recommenders deliver a superb and detailed portrayal of yourself. Be mindful that January and February... -
3 Tips to Improve HDR Efficiency for CRISPR Editing in Human Cells
TypeBlog Post...it is incorporated. We were surprised to see how rapidly the efficiency of mutation incorporation falls...contains the intended sequence changes to be incorporated into the edited genome. In mammalian cells such...these two strategies allows you to specifically incorporate homo- or heterozygous mutations. 1. Increasing...therefore strongly encourage any researcher to incorporate CRISPR/Cas-blocking mutations at both targeted... our frustration) in many cases we only saw incorporation of the blocking mutation. Despite the fact that... away from the cut site were not efficiently incorporated during HDR. We decided to systematically characterize...only about 30 bp, it is no longer feasible to incorporate mutations without screening thousands of clones... -
Quick Guide to Near-Infrared Fluorescent Proteins
TypeBlog Post...molecular brightness (Table 1). NIR reporters and biosensors NIR reporters can tell us about cellular events..., D. & Michnick, S.W. An infrared reporter to detect spatiotemporal dynamics of protein-protein interactions...FPs) were derived from natural photoreceptors incorporating biliverdin IXα (BV) autocatalytically. Fortunately...should be supplied (Baloban et al., 2017). It is important to consider effective brightness of NIR FPs in... a cell and depends on the efficiency of BV incorporation by an apoprotein and protein expression level...higher apparent cellular brightness than previously reported widely used dimeric iRFPs (Table 1). In general...Oliinyk et al., 2019). They were engineered to incorporate BV. Their advantages are their small size (17... -
Gendered Innovations: Why Does Sex of the Cell Matter?
TypeBlog Post...emphasizes that reporting the sex of cells and tissues—when that information is reliable—is important to ensuring...Universities need to incorporate these methods into their curricula. Corporations need to integrate these...Governments, universities, and, increasingly, corporations in the US and Western Europe have taken three...into research. Why are sex and gender analysis important? Doing research wrong costs lives and money. According...positive, the analysis did find 263,000 more osteoporotic fractures. The overall economic savings to the... goal of gendered innovations It is crucially important to get the research right from the beginning. ...unique to male stem cells. Nor did they detect important differences in function between male and female... -
Hot Plasmids April 2018 - Protein Degradation, Nanoscopy, FIRE-Cas9, and Yeast Expression Tools
TypeBlog Post...expression, sometimes incorporating chemically inducible approaches to provide temporal control, this new ...random variation, or noise, in reporter expression. To do so, one places reporter expression under the control...expression of a reporter protein, it is possible to change the amount of noise in reporter gene expression...can deliver these components together using electroporation. In these bulk cell experiments, Clift et al...no stress or damage upon TRIM-Away, and the electroporated cells re-adhered to the culture plate within...deliver the target antibody. TRIM-Away reagents and reporters are available from Addgene - we hope they will...lab has deposited plasmids encoding fluorescent reporters and other proteins controlled by GAL4 and ZIF268... -
Plasmids 101: FLEx Vectors
TypeBlog Post...gene. Spatio-temporal expression of optogenetic reporters: Since most optogenetic reporters are delivered...invert the reporter and GeneR fragment. Then the second recombination event would excise the reporter from ...transduced reporter gene. Generating a FLEx switch to control expression of an optogenetic reporter would ...Cre-dependent reporter expression, the potential for background expression of the reporter will be greatly...shown in Figure 2, the cassette should contain a reporter DNA sequence (such as GFP) in the sense orientation...the two pairs of target sites. By flanking the reporter with one loxP site and one lox511 site in the ...would ensure that the reporter remains silent until a cell or tissue-specific Cre is provided. Since the FLEx... -
How to Design Your gRNA for CRISPR Genome Editing
TypeBlog Post...sequence are important considerations for designing your gRNAs. For indels, it's not so important what location...nom. Me love gRNAs! Comic by Maya Kostman. Important considerations before you start an experiment ...tool among them. While this seems obvious, it is important to remember that the same is true when designing...location in the gene you target, but it is important that your gRNA sequence is designed to be highly active...CRISPRi, these considerations are of roughly equal importance (target should be near the TSS but you can worry...from). Finally, for HDR, location is much more important because you have to target within ~30 nt of your...gRNA with an optimized sequence is of primary importance (more on this below). Created with BioRender.com... -
Plasmids 101: Multicistronic Vectors
TypeBlog Post...along with a reporter, such as a fluorescence gene or a resistance gene. These reporters allow you to ...sometimes advantageous, particularly when only a portion of the plasmid is packaged for viral delivery, ...expression levels between two or more genes is important. How do multicistronic vectors work? Scientists...single transcript. The two strategies most widely incorporated into plasmids for research purposes are described... using IRES elements. Further, scientists have reported lower expression of the downstream cistron due...eukaryotic cells, and, although some scientists report close to 100% cleavage with some of these peptides...that already has the multicistronic element and reporter cloned in. In these plasmids you would simply ... -
AAV Titers: Where Do They Come From And What Do They Mean?
TypeBlog Post... people usually report? Addgene and other viral vector manufacturing facilities report the physical titer... characteristics give us information on three important factors. What can AAV titers tell you? Physical...cell transduction assays. Wild-type AAV2 has been reported to have a near-perfect physical-to-infectious ... However, for recombinant AAV2, the same study reported a physical-to-infectious particle ratio of 50:...(“full”) (Zeltner et al., 2010), while another reported that only 20% of recombinant AAV2 particles were...vector preparation protocol, and we are pleased to report that over 95% of viral capsids in representative...dosing purposes in preclinical studies, it is important to understand what these values mean and how they... -
Plasmids 101: Control Plasmids
TypeBlog Post...of the independent variable you are testing. Importantly, they demonstrate the functionality of your experimental... experimental system and help identify opportunities for troubleshooting or optimization within your experiment...non-independent variables. To illustrate the importance of control plasmids, we will take a step beyond...may be a plasmid that constitutively expresses a reporter protein (e.g., GFP or luciferase) that is either... well of your cells. Regardless, the amount of reporter protein activity correlates to both the amount...living, viable cells. This result presents an opportunity for troubleshooting and optimization of the transfection... transfection reagents and process. It is important to note that optimal experimental conditions, including... -
Addgene’s Viral Service - Why Virus? Why Now?
TypeBlog Post...milestone. Viral service supporters We would like to thank the companies who supported this project with funding...and be ready to provide extensive logistics support for export and shipping on dry ice. The Business and...seed money was used to build a completely self-supporting service so that Addgene is not dependent on grants... in accelerating the Viral Service. With their support we are able to provide faster access to a larger...number of viral preps. Visit our Sponsors and Supporters Page Thank you to all of our generous, sharing...Addgene repository. Without these scientists and the support of their technology transfer offices, Addgene would...Jonathan Ting’s lab from the Allen Institute have supported the AAV initiative from the beginning and have... -
The Addgene Antibody Data Hub
TypeBlog Post...Antibody Data Hub. Data reports Each antibody submission is used to create a data report, which contains experimental...that information in the report as well. Figure 3: An antibody data report, showing the results of ...methods section for the data report shown in Figure 3. At the end of the report, you’ll find a results section... the data report shown in Figure 3. The antibody is rated "Pass". Curating the data reports Just like... page of the Addgene Data Hub. Antibody data reports can be searched and sorted by gene/target, name...information is then formatted and published as a report with a DOI for citation. If you’re considering ... -
Making Career Development an Integral Part of the Science Training System
TypeBlog Post...Career Development; a Stakeholder Workshop Report.” The report enabled the development of priorities for...programs are still doing an inadequate job of supporting science trainee transition into diverse, successful...amplified by systemic biases which, in the absence of support and structure, prevent science from attracting ...Development Hub Workshop Attendees and their 2021 report. Image from pd|hub. To begin to tackle these... of Massachusetts Medical School. It has been supported by a Steering Committee and working groups with...the findings were captured in this just released report “Enhancing Dissemination of Evidence-Based Models...Institute of General Medical Sciences (NIGMS) will support the development and implementation of collections... -
Building Global Connections with the International Mentorship Program USA-EUROPE
TypeBlog Post... and logistic support to increase the chances of success, but also nurture a supportive and protective...Instead, talent should be realized through equal opportunities for all regardless of origin. It is therefore...first-hand vision of the challenges and career opportunities that truly exist outside their immediate frontiers...mentee, where (s)he feels truly understood and supported, and can more fully identify with and be inspired...the beginning, I registered at IMP to have an opportunity to improve my networking with mentors who are...how to write cover letters, and has suggested important things to emphasize in order to be selected in...“IMP showed me a wide range of professional opportunities that are outside of our typical boundaries, ... -
Fluorescent Proteins 101: Visualizing Subcellular Structures & Organelles
TypeBlog Post...can be gained by understanding the location and transport of the protein within a living cell. Analyzing...Analyzing differences in protein localization and transport between healthy and diseased states can also provide...endoplasmid reticulum, modified in the golgi and then transported in vesicles via the secretory pathway to their...the end of their life cycle, proteins may be transported through the vesicles to the lysosome where they...Once you've created your fusion protein, it's important to validate its function relative to the untagged... of an organelle or involved in intracellular transport pathways, it is possible to map the internal organization...used markers for major mammalian organelles and transport pathways. It is best, though not always possible... -
Technical Design of a Western Blot
TypeBlog Post...variety of pore sizes. If your protein of interest is small, you may want to consider a smaller pore size (...primary antibody required. Reporters There are two main classes of reporters: chemiluminescence enzymes...protocol and protocol video! Antibodies The most important part of your western is your antibodies, particularly...you need a specific conjugate, and changing your reporter molecule requires either re-conjugating or re-...-purchasing your primary antibody with the new reporter. Finally, each new conjugate will have to be tested... weight proteins Long running time It’s important to note that Bis and Tris gels run proteins in...high protein retention. It comes in a variety of pore sizes and protein capacity, allowing you to optimize... -
Scientific Sharing in the Time of COVID-19: Databases and Resources
TypeBlog Post... world. To support this pace of research, new, centralized databases and centers to support the sharing...like here at Addgene!), and new databases are supporting their access to useful resources during this ...information about reagents, funding and volunteer opportunities, protocols, publications, and more to accelerate...the COVID-19 Resources Canada site. To date, the portal has recruited 3,218 Canadian volunteers, and features...COVID-19 related research projects, funding opportunities, and community response initiatives. Accelerate...research Universities have also been stepping up to support COVID-19 research. The new Center for COVID-19 ...Preparedness (CCRP2) at Rutgers University not only supports collaborations at Rutgers, but also lists data... -
RNA Interference in Plant Biology: New Tools for an Old Favorite
TypeBlog Post...fluorescent reporter, directly couple silencing of your gene target to silencing of the reporter. This is...application of long or short dsRNA. However, it is important to note that naked and unmodified dsRNA elicits... Together, this excellent work highlights the importance of using orthogonal methods to confidently conclude...fusing your gene target sequence to a fluorescent reporter sequence (discussed below). Examples of these ...genome, whereas shRNAs are exogenously derived. Importantly, instead of the assortment of siRNAs generated...contains a target sequence against the fluorescent reporter in an inverted repeat orientation. Therefore, ...sequence against your gene(s) of interest and the reporter. This approach greatly enhances experimental throughput... -
Finding Your Science Policy Path
TypeBlog Post...governments. Most communities have opportunities to join commissions or boards, support ballot initiatives, deliver...are ways you can hone important skills for the profession. One of the most important skill sets for a science...success in a science policy career with many opportunities to pivot and grow along the way. That’s what...fellowships, volunteer positions, and part-time opportunities. But even through all my roles and my current... of the who, what, where, when, why and most importantly, how, of a science policy career. Who does science...science policy? I would argue that this is the most important question that you can explore. The answer will...advice for you and your next steps. This advice is important, but must be taken through your own personal lens...