...workplace, and pursue leadership opportunities at the local, institutional, regional, and national levels. These... first at Brigham and Women’s Hospital/Harvard Medical School and then Boston University (institutions...issues faced by postdocs and institutions are not local but are national issues. I applied and was successfully...other people mostly online and on monthly phone calls. This has its challenges as meeting in person is...Chair then stepped down, I became interim Chair automatically as it was an understood expectation of any Vice-Chair...

...and solutions to research problems. This movement calls itself the Open Science movement and is part of ...and the reagents are too expensive to make it practical - but we have members looking at how to re-use...don’t throw old equipment away until you ask your local makerspace if they want to hack new life into it...somewhere near you. The place to start looking for your local space is the Hackerspace website. Markerspaces differ...fun with technology, so feel free to contact your local space and get involved! Derek Jacoby is a PhD ...

...different questions than those on the prep calls, but often the prep calls provide good ideas for interesting ...possibly with an organizer) should set up a 30 minute call with each panelist to get to know their style and...better at remembering these things on a relaxed phone call than when first confronted on a stage in front of...long introductions. The moderator asks questions, calling on 1-2 panelist for each question in a way that...

... more than 10 modules often rely on two-step hierarchical approaches using different Type IIS restriction...stability, and buffer compatibility have placed practical limits on single- or two-step assemblies. We have... While T4 DNA Ligase, the mainstay of most biotechnological cloning efforts for over 50 years, prefers... plate Fidelity of assembly (percent correct) Calculated colony totals Per 2 μl assembly reaction Per...outgrowth volumes described in Figure 3, with calculated yields from entire outgrowth built from 2 µl ...

... molecular biology-grade agarose containing physiological concentrations of salts and glucose. If we circled...fluorescent protein), fluorescence microscopy can theoretically be used to select specific wells from the 48... terms of growth rates, morphology or other physiological properties. An advantage of this technique is...properties, either initially, or using secondary biological screens. For example, for cancer researchers,...Neurobiology and Anatomy at the University of Maryland Medical School in Baltimore. She is particularly interested...

...BV510 and AmCyan have almost identical emission spectra but may be called BV510 on one cytometer and AmCyan...for one another. This overlap in spectra is often called "spillover." For example, if your flow cytometer...shift towards the X axis to become a perpendicular vertical line.     In general, most spillover between samples...research technician in Arlene Sharpe's lab at Harvard Medical School.   More resources on the Addgene blog Introduction...

...the right fit for you without the financial and logistical stress that accompanies traveling for campus ...Visits After receiving applications, universities typically invite a select group of students to continue ...and each other. Before COVID-19, financial and logistical obstacles could make touring potential programs...in-person interviews for a variety of reasons (logistical, financial, etc…) now have options to fully engage...format of interview from a travel day to an online call in response to challenges on their end. Though the...

...Preger et al., 2020. Preger et al., Journal of Biological Chemistry 2020. https://doi.org/10.1074/jbc.RA120.01289...research, which required protein compartments to co-localize multiple enzymes in a reaction pathway, the MPyV...expression selectively in Parvalbumin containing cortical interneurons. We now provide ready-to-use AAV ...vectors expressing GCaMP6f with this promoter. Image calcium transients in PV interneurons with AAV-S5E2-GCaMP6f...terminal providing better signal to noise for imaging calcium dynamics in fine structures. We have expanded our...

...introducing the delivery method in 2014, Manuel Goncalves lab report that AdV-mediated transduction of gRNA...Gonçalves and his colleagues at Leiden University Medical Center are now available at Addgene along with ...and very efficient introduction of DNA into therapeutically relevant, non-transformed mammalian cells. ...and used AdV vectors to edit genes in several biological systems. It has been used for instance in inherited...Viral vectors for gene therapy: Current state and clinical perspectives. Biochemistry Moscow 81:700–708 ....

...in-house protocol. DNA yields for this procedure are typically in excess of 100 ng/ul, more than enough DNA to...be viewed more as a guideline, since they are typically based on the copy number of the empty plasmid ...improved plasmid DNA yield. Culture conditions play a critical role in plasmid DNA yield. Different plasmids ...are generally, but not perfectly, correlated. Typical growth times for high copy plasmids in standard...miniprepping at various time points. Measuring the optical density of the culture can indicate the number ...

... of these techniques have in common? They all typically require secondary antibodies, frequently of the...are derived from an unusual type of IgG antibody called a heavy chain antibody (HCab), which are unique...different species, nanobodies are useful for colocalization labeling of up to 3 targets (Pleiner et al ...desired fluorophores. In figure 4B below, triple colocalization is possible even when staining cells with three...multi-color staining workflow yields similar localization patterns as cells stained for one target (compare...

... science teachers at our school put out the call to local parents who would feel comfortable speaking ...parents who can answer some of the surprisingly technical questions elementary students can ask. It doesn... dangers of people using the technology in an unethical manner, but I also tried not to be an alarmist...fields in biology, are moving so quickly that it’s basically impossible for textbooks to keep up. Even the ...

... and ways to navigate it, we’ll search for the calcium biosensor, GCaMP, and click through to the plasmid...distribution. If your experiment involves delivering a calcium biosensor using AAV, you might decide you’d really...? The available filtering options now change dynamically based on the type of material and specific search..., followed by a convenient table of plasmids specifically designed for this purpose: Still stumped? Ask...results at all, let us know! Contact our scientific technical support at [email protected] and let us help.  ...

...Ample distribution channels both digitally and physically. Easy to incorporate visuals if needed. Traditional...means of scicomm. Digital or print distribution.  Typically limited to a single page of information. Primarily...levels and interaction. Can bring in things to be physically interacted with. Public speaking is a common ...internet access. High probability of real-time technical difficulties Exhibits Medium to high Audience...exhibit). No remote access; limited occupancy. Typically needs to be staffed. Mixed Media Medium to...

...especially in the sciences and by extension to scientifically oriented nonprofits. Like Deb, I worked for...available through not-for-profits, universities, local and national government offices, schools, healthcare... arts? Do you find satisfaction in helping your local food shelf raise money? Do you have experience with... as a grant writer and/or grant reviewer with a local nonprofit that aligns with your passion to get grant...Join the Grant Professionals Association (GPA)  local GPA Chapters, and the National Grants Management...

...the Janelia Research Campus of the Howard Hughes Medical Institute. Are you currently on or planning to ...rejection of a model and no clear alternatives. Typically, a research vision is broken up into three aims... basis of a single grant. Either way, think realistically about the time and resources required to feasibly... aims in more depth in the remaining time. The typical chalk talk structure is: What is the big question...the Janelia Research Campus of the Howard Hughes Medical Institute. His interests include the quality control...

... this post I’d like to delve deeper and look specifically at the characteristics and capabilities an ELN...facilitate true reproducibility. The following are critical: (1) connectivity to other data sources used in...likely stored in institutional and lab servers specifically designed to hold large amounts of data. Although...group’s research – it makes no sense and is not practical to move all this data in bulk into a lightweight...lightweight tool like an ELN. It is nevertheless critically important to be able to conveniently reference specific...

...molecules, forming a “handcuff” structure that physically blocks replication machinery when plasmid copy...It is also worth noting that plasmids can be symmetrically incompatible, meaning that both plasmids are... from the cell with the same probability, or asymmetrical, where one plasmid lost from the cell at a higher...deal with the more sinister plasmids of the microbiological world. (Want to know more about these plasmid...high copy incompatible plasmids that lead to asymmetrical plasmid loss of the usually large, low copy ...

...JAT, Callaway EM (2007) Monosynaptic Restriction of Transsynaptic Tracing from Single, Genetically Targeted...Why pseudotype? Lentiviruses and retroviruses typically target immune cells with their wild-type envelopes...out this review article published in Reviews in Medical Virology (Li et a., 2018).   Figure 2:...6178.1997 Wickersham IR, Finke S, Conzelmann K-K, Callaway EM (2006) Retrograde neuronal tracing with a deletion-mutant...

...studying biochemistry at the University of Southern California. He is also an intern at the Yang Lab of the ...transposon-encoded CRISPR/Cas systems in bacteria, specifically on optimizing strains with low efficiency.   ...of Sciences. He joined Shanghai Institutes for Biological Sciences in 2000 and became a professor in Synthetic...reprogramming of cells to understand the genetic and biochemical mechanisms behind their distinct phenotypes and...generation of engineered cells for green chemistry and biomedical applications.   Yiwen Zhang entered the Sheng...

...production of viral vectors typically involves shuttle vectors, also sometimes called transfer vectors. The ...antibiotic works. Another example is Zeocin, a DNA intercalator, which affects all kingdoms of life. Of course...host range plasmids shuttle vectors? While it’s technically accurate to say that shuttle vectors have a “... “broad host range,” these terms are typically used in different (though overlapping) ways. Broad host...

...product purity of 66%. Pro tip! Abasic sites are also called apurinic or apyrimidinic sites and abbreviated ...are the same protein!) (Chen et al., 2024). They called their initial editor AXBE, for A → X base editor...in a G → Y (Y = T or C) base editor, which they called glycosylase-based guanine base editor (gGBE) (Tong... Therapeutics team has deposited their editors, called gTBE and gCBE, with Addgene. gTBE can introduce...site A DNA site with the nucleobase removed. Also called an apurinic / apyrimidinic (AP) site. UNG / UDG...

...In Addgene’s early days, we would ‘spot-check’ critical regions of plasmids deposited with us via Sanger...development teams, frequent updates allowed us to scale our lab work as we grew. As the number of deposited...us to switch between executing our pipeline on a local computer or in a cloud environment. Given the throughput...jumping back and forth between running the pipeline locally and in the cloud. Creating a bioinformatics workflow...script file (containing our new process) on our local computer. Eventually, our workflow would handle ...

...experimental design and its potential for clinical applications. Specifically, efficacy rates of Cas7-11 were reported...current knowledge. The Abudayyeh-Gootenberg lab (also called the AbuGoot lab), a joint lab run by Omar Abudayyeh...originally suspected that this new Cas7-11 might be phylogenetically classified as the more complex Class I systems...switched up the serine integrase for one which can enzymatically take a genetic sequence and integrate it into...

...expression limiting regulatory elements or use a non-canonical code (Gustafsson et al., 2004), such as Hoogsten...recognize multiple synonymous codons) may actually be critical for determining the rate of translation. The ribosome...10.1101/gr.139758.112 Mauro VP, Chappell SA (2014) A critical analysis of codon optimization in human therapeutics... GluCl ion channel genes for mammalian cells dramatically improves expression levels. Journal of Neuroscience...synthetic? How Addgene;s dataset reveals trends in biological innovation Resources on Addgene.org Check out...

...regions and allocate specific functions to them. Electrical recordings and lesioning studies have been essential...within the LC of mice. When those neurons are chemogenetically activated, norepinephrine release causes a...an enhanced propensity for movement and activity called locomotor sensitization. This enhanced movement...responsible for the antidepressant effect. To specifically activate the Drd1 neurons, they injected AAV2... "Hypothalamic CNTF volume transmission shapes cortical noradrenergic excitability upon acute stress."...

...post was updated on Jul 27, 2020. CRISPR, and specifically Cas9 from S. pyogenes (SpCas9), is truly an ...specified by the crRNA sequence, it adopts an enzymatically “active” state rather than reverting to an inactive...propensity of Cas13a to bind and cleave RNA non-specifically. The ability of dCas13a to bind target RNA without...modulating splicing, and regulating expression of specifically targeted RNAs. Finally (and perhaps most interesting...observed shRNA off-targets. They subsequently used catalytically inactive dLwaCas13a as a programmable RNA-binding...

...FRET during a journal club, my guitarist brain automatically thought about the raised element found on the...interactions in living cells using a microscope. Historically, molecular interactions were detected by indirect...know or interact with each other. FRET reduced the scale of our perception about molecular interactions. ...Periasamy defined FRET as "a distance-dependent physical process by which energy is transferred non-radiatively...FRET) microscopy imaging of live cell protein localizations. J Cell Biol. 2003 Mar 3;160(5):629-33. ...

... RT-qPCR tests to diagnose COVID-19 and some serological tests to determine past exposure. However, the...rapid detection.  Highlights: Cas13a is not catalytically active in the presence of two or more mismatches... copies RNA targets in a 40 min incubation. A scalable, easy-to-deploy, protocol for Cas13-based detection...available and less expensive. Instead of using the typical PCR machines, the authors use affordable, field-ready..., Acosta-Alvear, D., & Wilson, M. Z. (2020). A Scalable, Easy-to-Deploy, Protocol for Cas13-Based Detection...

...set of transferable skills, coupled with your technical skills, can open new career opportunities for ...conflict can also be good. In science, a reviewer’s critical comments on your manuscript is a form of (hopefully...scientist. And some amount of conflict is actually critical in the business world. Companies and organizations...aspects of customer support each team “owns” is critical for day-to-day operations and keeping our customers...especially people with whom you strongly disagree, is a critical component of conflict resolution. Having empathy...

...insights into the complicated physical organization of the brain and other biological systems. Interested in ...deletions are defined using 2 loxP variants (only identical loxP sites can mediate recombination.) The deletions... preventing further recombination. Brainbow-1.1 scales up the system to four fluorescent proteins, with...homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells. Snippert HJ, van der...

...target specific tissues, even when administered systemically, without causing disease? Recombinant adeno-...genomic structure of AAV2, so they are formally called rAAV2/1, rAAV2/2, and so on, although we often ...in-stock viral prep service.  AAV5 AAV5 is the most genetically distinct AAV. Though AAV5 is one of many serotypes...on the AAV capsid. Molecular Therapy. Methods & Clinical Development, 26, 331–342. https://doi.org/10.1016...is required for gene transfer. The Journal of Biological Chemistry, 276(23), 20610–20616. https://doi....

...without their problems, but much genetic and physiological data have been accumulated over the years using...check out our Mouse Modeling blog posts: Part 1: Genetically Engineered Mice and Part 2: Breeding and Crossing...into the oviduct of pregnant females, a technique called Genome editing via Oviductal Nucleic Acids Delivery...small mutations with HDR, when it comes to larger scale genome editing, such as replacement of a mouse gene...nice to your microinjectionist, who has the strategically important job of delivering the payload! Last...

...lab, or tissue culture as it is commonly called, is a critical tool for many scientists. Mammalian cells...AAV preparation. For detailed instructions and calculations on how to prepare a plasmid stock used to generate...cells provide scientists with the means to study biological processes on the cellular level instead of having...

...completing her PhD in Chemistry and Chemical Engineering with Harry Gray at Caltech, Crystal moved on to a Postdoc...Investigator III at Novartis, a multinational pharmaceutical company based in Switzerland. After completing...scientific teams at the Novartis Institutes for BioMedical Research to develop novel biologics for the treatment...

...Addgene podcast, we sit down with Alex Chavez, a Clinical Fellow in Pathology at Masachusetts General Hospital...Church and Collins labs at the Wyss institute for Biologically Inspired Engineering. Alex is the first MD PhD...and explains why he loved the first 2 years of medical school. 7:05 - 10:55 : Alex's focus during graduate...postdoc 12:01 - 13:21 : The Wyss Institute for Biologically Inspired Engineering and why Alex decided to...

...use as a genetic engineering tool is frequently called recombineering - short for homologous recombination-mediated...insert site (Figure 2, top).  These primers are typically ~70 nucleotides long (20 nucleotides that anneal...fragments, such as gene replacements and tags.  The typical frequency of recombinants is 1 positive clone out...efficiency than the other and you’ll find out empirically which is the lagging strand. ssDNA substrate ...below).  Controlling expression of Red proteins is critical for minimizing the toxic effects of Gam expression...

...networks in biological systems. NGS will also play significantly into the realm of clinical therapeutic...validation process, also known as CRISPR genotyping, is critical to demonstrating causal relationships between ... forms of off-target analysis in order to meet clinical approval. It is key to maintain a consistent set...the detection of KRAS mutations: diagnostic and clinical implications." The Journal of Molecular Diagnostics...

... is necessary for bacterial transformation of chemically and electrocompetent cells is minimal (picogram...nucleic acids (genomic DNA and RNA), proteins and chemical contaminants that have been used during the purification... preparations for this technique must be larger scale, to allow the extraction of enough plasmid to last...addition to being free of any nucleic acid and chemical contaminants, these DNA samples must also be entirely...

...in Plasmids 101, tags can help you determine localization of a protein of interest, purify it, or determine...major caveat - a tag may interfere with protein localization and/or function, so each tagged protein must...proteins found that 20% of tagged proteins did not localize similarly to their untagged counterparts. These...fluorescent-protein tagging show high correlation for protein localization in mammalian cells." Nature methods 10.4 (2013...