...of the opportunities you’re provided throughout your PhD program.   Find people who will support your ...had a wonderfully supportive mentor and graduate office. I cannot stress how important they were to my ...experimenting with the career and networking opportunities available to me as a graduate student culminated...events for adults and professional development opportunities for researchers interested in connecting with...they had my back. If your advisors are not as supportive of your career goals, find people who are. I ...as types of experiments. I searched for new opportunities with an open mind and open ears. Everyone I ...mentioned, my graduate office staff was amazingly supportive. I talked to them about my interests and my ideas...

...everywhere can access plasmids despite the evolving export/import rules. Our shipping and lab teams continue to...entire supportive Board, we have a robust committee process in place and continue to incorporate additional...all of the success we have achieved and the opportunities we have for the future. At Addgene we will never... The success of these projects allows us to support more new projects. Diversification is happening... quality and success. Customer and technical support Our customer and technical service brands continue...sharing, but we don’t have to worry about getting support from depositors. We regularly hear how much they...blessed with a large community of scientists who support us by depositing, requesting and spreading the ...

...benefit from some Addgene support? Please get in touch, as we aim to support a couple of events a year...across Europe get introduced to the cultures and opportunities available to them at university. At a Freshers... into university life. The fairs are also an opportunity for the societies to promote their upcoming events...biological societies that Addgene has had the opportunity to work with at Freshers Fairs in the past year...undergraduates and PhD students so that they can help support each other and provide advice. The society organizes...involved with. The society provides talks and career support for a range of different scientific disciplines... in its first year. The society facilitates opportunities to learn about Synthetic Biology and exciting...

...et al. Cell. 2015. PubMed PMID: 26627737 pORTMAGE: A portable method for bacterial genome engineering ...-ome using a quick, affordable, and ubiquitous reporter assay. In their recent publication, Kroeze et ...their enhanced TANGO arrestin recruitment assay incorporating 314 codon-optimized GPCR sequences for human...MAGE uses recombineering (2) to simultaneously incorporate multiple single-strand DNA (ssDNA) oligonucleotides...so-called “genomically recoded organism” (3) but its portability to other bacterial strains remains seriously ...bacterial species (4). This set of plasmids (dubbed pORTMAGE) expresses the λ Red recombinase enzymes, as well...can be used in a broad range of strains. Thus, pORTMAGE simultaneously allows genome editing and mutant...

...plasmids, are export-controlled materials in the USA, where Addgene is located.  The export license, if ...requestors. We also offer export license applications for VSV and SARS-COV-1 plasmids. Export control license ...Addgene is excited to announce we now offer an export license application for rabies materials! Some ...license process    An export control license, once approved, allows Addgene to ship plasmids or viral vectors...the applicable research project when purchasing export-controlled materials. Addgene will then apply for...process.   Due to recent regulation changes, an export control license for rabies materials may be required...required for others. You will be notified if an export license is not required for your country after ...

...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...

...Fluorecent Proteins in Nano Lanterns Reporter gene assays A reporter gene, such as luciferase, usually serves... is also the most commonly used bioluminescent reporter. This beetle emits a yellow-green light with a...several studies utilized luciferase as a genetic reporter in plant and mammalian cells. Luciferase assays...available to choose from) is now a common feature in reporter plasmids.  Learn How Luciferase Can Be Used In...transcription within cells, where detection of the reporter protein or its enzymatic activity is measured....expression can be determined by detection of the reporter in a specific assay, which ideally would have .... Production of photons by this bioluminescent reporter occurs slower than fluorescent-based methods, ...

...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...

...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...

...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...

...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...

...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...

...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...

...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...

...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 ...

... 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...

...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...

...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...

...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 ...

...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...

... 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, ...

...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...

...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...

... 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...

...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...

...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...

...fundamentally the reporter construct monitors the expression of a nearby gene. The reporter gene serves as... screens to introduce reporters throughout an organism’s genome. These reporters have historically been...stimulate the expression of the reporter. The system is made up of a reporter gene flanked by a minimal promoter...promoter – all rely on the random integration of a reporter construct in close proximity to an endogenous ...example, if green fluorescent protein (GFP) was the reporter gene and green cells were observed in the kidneys... for with an uncharacterized gene. The actual reporter construct can be something phenotypic (such as...promoter. The system is composed of a promoter-less reporter which needs to integrate within an exon of a gene...

...External collaborations are becoming increasingly important as more complex approaches to scientific questions...collaborations and partnerships are also very important in a non-research context, especially when considering...expand from there. Start small: Collaboration opportunities within an organization but outside your comfort...graduate or postdoctoral work there are loads of opportunities to create communities, platforms, and to be ...Relationships” below). Expand: Collaboration opportunities outside an organization The next step is to ...out as a student, people tend to respond in a supportive way and are willing to help because you are not...work. Get sponsors for a training weekend for a sports club, find speakers for a meetup or recruit volunteers...

... transformation. Choose good reporter genes There are many great reporter genes available and which one...half-lives then GUS and are therefore good temporal reporters to measure promoter activity, for example...the T-DNA region with our gene of interest or a reporter gene so that the transformation and gene expression...different treatments on transformation success. Some reporter gene examples are below. The GUS gene (β-glucuronidase...histochemical GUS-staining is a very useful visual reporter (Figure 2). The GUS enzyme cleaves a substrate...transformation success. Luciferases are great quantitative reporters and use a substrate to produce bioluminescence...Agrobacterium. This is especially problematic with reporter genes as you might observe expression from Agrobacterium...

...as to where readers may locate data supporting the research reported in PLOS publications. The policy was...published data. The importance of data sharing The reason this topic is important to my lab is three-fold...available for anonymous request.  This means that supporting datasets must be logically annotated and arranged...work this may entail.  So if access to data is important, what data should be included? We should recognize...is calculated and placed in the legend. Widely reported difficulties in reproducing experimental data ... in scope. For images, the number of examples supporting the published material is a factor of practicality...over time such that prior formats are no longer supported/readable. Is the author or their host institution...

...believe that communication is the single most important skill that scientists need to succeed in their... always recognized and valued for its immense importance, it may well be what determines whether you get...wasted if you cannot convince others that it is important and relevant to them.   When you think about it.... Treat every interaction in your work as an opportunity to practice communicating.  Excellent scientists...you will also have to explain why the work is important and how it should change how the members of your...education and policy and to draw attention and support to work in their field. It can be hugely beneficial...ComSciCon, our primary mission is to strengthen and support the network of graduate students who are passionate...

...a UAS-driven luciferase reporter construct. In the absence of GFP, no reporter output was observed in ... in vivo system, Tang et al. electroporated T-DDOGs, GFP, and a reporter construct into the mouse retina...activated the reporter gene TdTomato, whose expression was absent without electroporated GFP. They then...specific. When tested in retinal electroporation studies, DsRed reporter expression was induced in ~76% ...and Cre-DOG, respectively, repurpose popular GFP reporter lines for more sophisticated experimental manipulations...then tested T-DDOGs with two mouse GFP reporter lines; again, TdTomato was seen only in cells with GFP,...

...future career opportunities. Becoming involved is not too difficult and you can find opportunities around you...within their workplace, and pursue leadership opportunities at the local, institutional, regional, and national...As the Chair of the Advocacy Committee, I had to report to the Board of Directors and liaise with the Executive...accordingly. For example, the quarterly and annual reports I had to write on behalf of the Advocacy Committee...to overcome the personal frustration and submit reports in the format requested which in the long run was...invited to spend a weekend together, funded by supporters such as the Burroughs Wellcome Trust. These “...experienced and learnt. Opening doors to further opportunities  Having gained leadership skills and advocacy...

...as a striking example of the applicability and importance of research in basic biology, medicine, epidemiology...of what you’re getting yourself into is really important.  As you’re talking to people, though, try to ...your time is valuable! Don’t jump on the first opportunity that comes your way. Think critically about whether... found that in the right environment and with supportive mentors, being open and vulnerable about my struggles...fundamental understanding of what you’re working on is important! Better than leaving yourself in the dark, but...have often provided me with much-needed emotional support during my journey. When you are hunting for labs...join, one of the vital things to look for is a supportive environment—it makes all the difference in a ...

... outcomes and the opportunity to make new discoveries. Provost notes the importance of acknowledging student...history. Interestingly, he also adds to the disproportionately large number of honorees to have trained ... and he wanted to provide the same kinds of opportunities for his students. But he also found that his...Bringing his research into the classroom. I had the opportunity to talk with Provost recently about his development...Undergraduate Research Experience (CURE). Having incorporated a discovery-based lab on planarian regeneration...interaction with other proteins.   It is important to note that research activities in a CURE must...with colleagues at other institutions to form a support network called the MDH CUREs Community (MCC) that...

...are removed and replaced by a single (non-viral) reporter gene (e.g. firefly luciferase or eGFP) flanked...necessary to direct minigenome replication and reporter gene transcription by the EBOV polymerase (reviewed...viral proteins are provided in trans (see Other Important Components of EBOV Minigenome System). However...negative sense RNA viruses (reviewed in 2, 3). Other important components of the Zaire ebola virus minigenome...the minigenome as a template to transcribe the reporter gene (e.g. firefly luciferase). The polymerase...start signal and initiates transcription of the reporter gene. The polymerase will elongate the mRNA until...target the viral polymerase. Technical tips It is important that all of these plasmids (those containing NP...

...however, offer you an opportunity to make a personal connection; the importance of which will be elaborated...what comes next. To that end, one of the most important things that you can do now is diversify. Read ... position! Conferences One of the single most important things you can do to plan for your career is to... Speaking of jobs, it seems like an obviously important experience, but far too many students and young...several per month, giving you a plethora of opportunities to explore! Again, attending one of these fairs...can search for jobs, post your CV, and, most importantly, manage your professional network. This is probably...box (most likely your lab, in this case). It is important to realize that you have to take responsibility...

...combination with that reporter, you can infer its presence through detecting the reporter's fluorescence. Another...labeling proteins is to fuse them with a fluorescent reporter, such as GFP or mCherry, through genetic modification...combination of antibody-fluorophores, fluorescent reporters, and/or dye stains used in your flow cytometry...excitation and emission wavelengths, it is also important to know the brightness of individual fluorophores...mentioned previously, you can also use fluorescent reporters, such as GFP, and fluorophore-conjugated antibodies...compare the excitation/emission spectra of your reporter with the ones from your fluorophore labels and...cells to express: CD45. Upon expression, CD45 is incorporated into the plasma membrane and can be detected...

...hypothesis. The results of experiments can help support or reject a hypothesis. Looking at the data, scientists...trust in scientists if the march highlights data supporting unwanted results (14). Some loss of trust, I ...science) they come from many different places supporting many different positions. It would be detrimental...going change depending on the number of people supporting it.  3) Linking scientists to one particular ...world. There is no requirement that marchers or supporters be scientists – on the contrary, anyone and everyone...the march as well as other science advocacy opportunities. Information for joining the march: March...satellite marches. Anyone can join the marches or support the cause. For science to be integrated into politics...

... the TIMER growth rate bioreporter. During protein maturation, the bioreporter spontaneously changes from..., and evidence suggests that it is incredibly important for nearly every aspect of life.” Unger began ... and in assays for measuring human serotonin transporter activity. Unger also plans to use the serotonin...permeability of water and nutrients and colonization opportunities for bacteria. To compare the artificial leaf..., Bernach plans to use a suite of fluorescent reporters. Bacteria containing unstable fluorescent proteins...artificial leaf, Bernach will use the CUSPER bioreporter such that fluorescent proteins expression is ...colonizer, labelled with a constitutively fluorescent reporter can also be used to track how one population responds...