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...This post was contributed by Doug Richardson, Director of the Harvard Center for Biological Imaging and...block out of focus fluorescence from reaching a detector or restricting fluorescence to specific excitation...light passes through a pinhole before reaching the detector. The pinhole is specifically placed to only allow...allow light from the focal plane to reach the detector and all light from above or below is excluded. ...the optics of a microscope before it reaches the detector. This means that two or more objects lying within...collected and projected back to a fast, sensitive detector such as a scientific CMOS camera. If plasma membrane...higher photon flux via the more efficient camera detector and lack of a light-reducing pinhole. By collecting...

....com. Mentors You may have an adviser, but do you have a mentor? There is a difference. A mentor is someone...that many graduate and post-doctoral programs lack any formal or mandatory professional development training... you may need to look a bit harder to find your mentor. This may be a connection you’ve made at a conference... Alternatively, it might even be a colleague. A mentor does not necessarily need to be someone a lot older...planning, has written a great blog on choosing good mentors for scientists. Continuing professional development...popular job site webpages. Importantly, find a good mentor that can share relevant experience and resources...

...Cytosine base editors with minimized off-target DNA editing The original cytosine base editor, BE3, induces...deamination. They generated these editors using the BE4max architecture. The new editors reported in this study ...the paper in Nature Biotechnology Find the base editor plasmids CRISPRi in Caulobacter crescentus ...ctrA, a gene encoding an essential cell cycle regulator. The lab expressed both dCas9 and the ctrA sgRNA...lab developed new fluorescently labeled piggyBac vectors to deliver uniform and sustained expression of ...editing by TET1 by co-delivering TET1 and protein factors for linking methylcytosine oxidation to DNA repair...

...place. As a curator of the thousands of amazing plasmid technologies contributed by our depositors, Addgene...plasmid" section Anytime you use a plasmid from the repository, we request that you cite it using both its Addgene...increases the visibility of both you and the original depositor. Our citations feature makes it so that, when ...reliability of our search results depends upon you, our depositors, giving us as much information as possible during...you to browse through the many plasmids in the repository, our scientists have curated a number of special...attention to the great variety of plasmids in the repository. And last, but certainly not least, we often ...Special Collections Pages Suggest Plasmids for the Repository Find Plasmids for Your Research  ...

... chimeric antigen receptors (CARs). CARs are a chimera of scFvs and T cell receptors and are expressed...portion of the receptors recognizes an antigen of interest while the intracellular T cell receptor portion of...domains of the selected scFvs are inserted into a vector encoding the antibody constant domain scaffold ..., and enzymes for cancer, autoimmune, and/or inflammatory therapeutic applications. Antibody fragments...cancer-related antigens, binding of the scFv part of the receptor can activate T cells to kill cancer cells. scFv...small size also allows for their delivery by viral vectors like AAV, which could be useful for delivering ...doi.org/10.1038/nbt.1791  Deal CE, Balazs AB (2015) Vectored antibody gene delivery for the prevention or treatment...

...papers or are brainstorming new technological innovations. Let’s take a look at three historical techniques...that save both time and money. Yet, knowledge of historical techniques comes in handy whether you are perusing...real-time PCR Southern blotting, named for its inventor Edward M. Southern (1), can be used to detect ...labeling it. The label can then be visualized by autoradiography and X-ray film for radioactively labeled probes...DNA technologies that are now commonplace in laboratories. As scientific technologies rapidly progress...throughout the world. Let us know your favorite "Historical Lab Technique" in the comments section below!...Coulson. "DNA sequencing with chain-terminating inhibitors." Proceedings of the National Academy of Sciences...

...in the specific repositories (Addgene, WormBase, Arabidopsis.info). These repositories will ensure that...post was contributed by Magdalena Julkowska, a postdoctoral researcher at KAUST, Saudi Arabia. From the...service that we, as scientists, provide for journal editors to help decide whether work is suitable for publication... submission to the subsequent journal.  As a postdoctoral researcher studying plant development and abiotic...the data can be deposited in the public on-line repository (like GitHub, Zenodo, Figshare, Triad) while ... and Records: The Royal Society Journal of the History of Science 69.3 (2015): 337-352. PubMed PMID: 26495581...protocols Find eBooks on plasmids, CRISPR, viral vectors, and more ...

...therefore we are not a party (signatory) to the MTAs for plasmids in our repository. Instead, we facilitate ...and Recipient (in Addgene parlance, the depositor and requestor) organizations. Over the last twenty years...with the number of requests and deposits in our repository. From 2004 to 2023, we facilitated 784,795 MTAs...MTAs per year.  The MTA Process at Addgene As a repository, Addgene doesn’t actually own the majority of...your organization agrees to let Addgene receive, store, and distribute your material under the specified...move forward with shipping the material to the requestor.  Why the MTA?  The MTA process can be a bit confusing...

...fully-equipped laboratory for citizen scientists. In addition to maintaining a laboratory to support both... of independent Do-it-Yourself (DIY) biology laboratories working towards enabling everyone to participate... set on top of filing cabinets, a rickety old incubator, a few pipettes, and some home-made equipment....Anyone can become a member of BosLab and previous laboratory experience is not required. Our members are skilled...have non-traditional scientists working in our laboratory! Don’t worry though, we make sure that everyone... became our bacterial transformation guru and instructor for our intro to biohacking courses. Our members...a co-organizer at BosLab, a community biotech laboratory in Somerville MA. Follow her on twitter at @ZebraElement...

...licensed distributor) allow you to quickly transfer a DNA sequence from the transposon vector to one of...encourage you to deposit your piggyBac™ transposon vectors with us to help us expand this useful resource....such as with signaling molecules or transcription factors, or when the gene is constitutively-active or a...edit of interest. With the larger piggyBac™ donor vector, it is possible to make the ITR and selection cassette...re-cutting. Transposase removal of the cassette later restores the target sequence. This allows researchers to...Transposase Read up on Genome Engineering with FLEx Vectors  Learn how Homology Directed Repair Is Used for...Genome Editing Resources on Addgene.org Find Vectors with the piggyBac™ Transposon Catch up on Everything...

... DNA sequence called the Fertility factor (F-factor). The F-factor is found on an episome, a piece of ... information on viral vectors, including transduction download our Viral Vectors 101 eBook. Bacterial ...fragmented DNA released into the environment. Various factors promote natural transformation in different bacteria...bacterial or eukaryotic cells via a virus or viral vector. One example are bacteriophages that attach to ...scientists commonly use phagemids, a DNA cloning vector that contains both bacteriophage and plasmid properties...

...reporters require a chemical substrate (and sometimes cofactors) to be provided and are generally dimmer than ...Kinetics of photon generation Slower Faster Cofactors/substrates Required Not required Signal strength...engineered Peak emission (nm) Size (kDa) Substrate Cofactors Secreted North American firefly (Photinus pyralis...AkaLumine ATP No Luciferase reporter assays for monitoring gene regulation Since firefly luciferase was ...expression analysis to investigate the effect of regulatory elements — such as promoters, enhancers, or untranslated...untranslated regions — on gene expression. The regulatory element is cloned upstream of the luciferase ...energy from an excited donor fluorophore to an acceptor fluorophore, which then emits it as light. Bioluminescence...

...entire mission is to enable that process. Our repository of plasmids, AAVs, antibodies, and data allows...technical skills, and experiences from thousands of laboratories that makes up our Help Center, blog, website... the blog originally started as a way to easily store and share in-depth answers to frequently asked questions...on a case-by-case basis, as many of our blog contributors primarily write in the areas in which they have...graphic designer upon request that the science communicator "just sketch a western blot out quickly." Final...and cited figures from papers or image sharing repositories. Then we format, do a final review to catch ...informative resources that answer questions our depositors and customers are asking. So how can you get ...

...contributed by Maaike Pols, PhD, a Developmental Editor at F1000Research.  With many changes taking place...their funding should also be published. All these factors will influence your choice of journals to submit...communities. The ARRIVE guidelines for the reporting of laboratory animals during in vivo research and the CARE ... are two sets of guidelines frequently used by editors. Providing all this information makes your manuscript...who the reviewers are). If the reviewers and the editor decide that your manuscript needs to be revised...the conclusions of your paper. This will help the editor handling your manuscript decide on whether your... is appropriate for that journal. Argue with an editor or reviewer over their decision. If you do object...

...proteins, such as human growth factor, vascular endothelial growth factor, and interleukin-6 (Witkiewicz... was contributed by Bárbara Pinho, a science communicator at the Portuguese science museum "Fábrica Centro...myofibroblasts, neuroendocrine, adipose, immune, inflammatory, angiogenic vascular, endothelial, and stromal... in increased expression of transforming growth factor-β signaling, possibly inducing the secretion of...seems to attack us by releasing mitogenic growth mediators and improving cancer cells proliferation.  How...enzymes and metabolism reprogramming aren’t the whole story. It’s essential for a tumor to spread and to resist... Pinho! Bárbara Pinho is currently a science communicator at the Portuguese science museum "Fábrica Centro...

...is a STEM career counselor, Director of Education for the Jackson Laboratory for Genomic Medicine and a... successful and sustainable biomedical research sector at our first Future of Research symposium. The ...compulsory internship to be taken in an industry, regulatory, or clinical environment. It remains contested...the overall structure of the biomedical research sector, and what a tenable workforce structure may look...ideal and productive workforce look like for the sector – how many jobs are there for STEM graduates and...strong supporter of increasing the number of postdoctoral scientists at University of Maryland. Paula... at Scientific Meetings from Addgene Executive Director Joanne Kamens Learn how to be a successful manager...

... be viral vector jokes and comics. Other new resources for researchers using viral vectors In addition...Addgene blog Download our Viral Vectors 101 eBook Find all of our viral vectors blog posts Learn more about...Starting today, our Twitter feed will be full of viral vector information. Each day, we’ll cover a different ...AddgeneGoesViral to find information on the basics of viral vectors, applications, and some new resources available...also investigates new strategies to improve viral vector production and data analysis. Our first paper came...

...in viral vectors, but easily fit in a TE. Transposons are also less likely than viral vectors to induce...DNA transposons are most commonly used in the laboratory for genome manipulation. When transposons are...transposon’s LTRs, similar to when packaging viral vectors.   Let’s take a closer look at three different ... in human cells at rates similar to retroviral vectors. The hyperactive version of the SB transposase,...efficiency in human cells similar to retroviral vectors. The hyperactive PB transposase (hyPB) is ~10-fold...transfer Transposons are an alternative to viral vector delivery of transgenes, like for iPSC reprogramming... piggyBac, which is a big advantage over viral vectors (~5 kB payload for AAV and ~8 kB payload for lentivirus...

...Hendel et al. 2015). Low detection rates mean the investigator can be relatively certain that their samples...off-target mutations are essential data points for investigators looking to definitively link genotype and phenotype...Targeted amplicon sequencing doesn’t tell the whole story Figure 1: Schematic representations of the...viable options for multiplex sequencing in many laboratories. Nevertheless, unbiased detection isn’t as sensitive...genome-wide unbiased methods. However, most investigators use single cell clones for in vitro CRISPR experiments... Hough. We additionally thank Monica Sentmanat, Victor Dillard, and Ayokunmi Ajetunmobi for their contributions... Hough is a Biochemistry PhD candidate in the laboratory of Prof Steve Jackson at the Gurdon Institute...

...nick, and monitor indel frequencies (Anzalone et al., 2019). Figure 6: Prime editors bound to ... Figure 4: Editing the PAM prevents the prime editor from re-engaging with DNA it has already edited...to pegRNAs drive editing (left), while prime editors bound to sgRNAs only drive nicking (right). Start ...opposite strand. Created with BioRender.com. MMR inhibitors PE4 and PE5: These approaches inhibit MMR to...inhibited. Created with BioRender.com. RNA protectors epegRNAs: epegRNAs include structured motifs...primer binding site (Nelson et al., 2022). PE7: This editor includes the fused RNA-binding protein La to protect...software, however. You may find it useful to ask a mentor what software tools they recommend — feel free ...