...deletions, and all possible base-to-base changes.   1. Plasmids 101: Transformation, Transduction, Bacterial...resources can now be found in one place: the Educational Resources page. So let’s get to it. Here’s the...Anti-CRISPR, 2in1 Cloning, Fluorescent Voltage Indicators, and Photoswitchable Proteins by Cary Valley,...protein size and shape, linker sequences, and fusion location, there are many choices to make. 2. Prime Editing...Resources at Addgene Browse the repository Find educational resources at Addgene Search for plasmids at Addgene...

...base to the other. Figure 1: Base transition editors catalyze purine-to-purine or pyrimidine-to-pyrimidine...engineering and protein delivery. Nature Communications, 8(1), 15790. https://doi.org/10.1038/ncomms15790...SpCas9 base editors with editing windows as small as 1-2 nucleotides. To reduce off-target effects associated...arrows indicate a conversion directly catalyzed by the base editor. Dotted red arrows indicate the change... point modifications. CRISPR base editing persists through multiple cell divisions, indicating that this...target modification in primary T cells, making them a promising tool for cell therapy applications. Dual...Base editing publication highlights In the table below, you'll find a list of publications we've highlighted...

...Zhang Lab's Nature Protocols Publication describing them here. References 1.Ran, F. Ann, et al. "Genome...

...2013 edition. 10 new developments in open science 1)      In February, the Office of Science and Technology...support for open access to peer-reviewed journal publications, although many don’t necessarily publish in ...

...P2A-PuroR for lentiviral expression. Figure 1: dCas9-p300 adds H3K27ac marks to histones in promoter...lentiviral transduction. Lysine-specific Demethylase 1 (LSD1) Tatjana Sauka-Spengler's lab has deposited ...epigenome (the sum of epigenetic modifications across the genome) has been implicated in disease states, and targeting...DNA sequences. Non-editing CRISPR applications direct catalytically dead dCas9 fused to a variety of epigenetic... the catalytic domain of p300 acetyltransferase increases levels of H3K27ac histone modification at specified...Preps for Epigenetic Modification Why use epigenetic modifiers? Epigenetic modification is certainly not ...Nature Communications 8: . https://doi.org/10.1038/ncomms16026 Find plasmids from this publication at Addgene...

...to western blotting  In a western, proteins are: (1) separated by size, (2) transferred to a membrane,...  (3) detected using antibodies.   Figure 1: Overview of the Western blot process. The sample,...post-translational modifications, quantify protein levels relative to standards, detect the cellular location of proteins...pull-down assays.  Westerns are divided into two categories, native and denaturing. In a native western, ...matrix, the blotting membrane, the gel, a copper cathode/iblot gel matrix, and filter paper. Pretty much...search, first pay attention to the validated applications listed on the vendor’s website and choose an...western and vice versa.  In addition to validated applications, pay close attention to the controls listed ...

...target cell line or organism (Presented in figure 1). More information on Agrobacterium-mediated transformation...mediated cloning of your gRNA of interest. Figure 1: Simplified schematic of agrobacterium-mediated transformation...current CRISPR research focuses on its potential applications for human medicine (Waltz, 2016), the potential...engineering, on the other hand, allows for targeted modification of known or suspected genes that regulate a ...model organisms like Arabidopsis and Medicago truncatula and several crop species including potato, corn...engineering using CRISPR, highlight the specific modifications to CRISPR machinery that allow for the use of...CRISPR principles). However, plant-specific modifications to commonly used CRISPR plasmids are necessary...

...synapses and deep-brain fiber photometry.  Figure 1: Characterization of iGluSnFR4 in mice. A) Scheme ...CycT1 or PolII-CTD antibodies and unlabeled MS in DLD-1 cells (colorectal cancer). Image courtesy of Brianna...Here's what you'll find in this post: Glutamate indicators with increased sensitivity and tailored deactivation...viral preps targeting interneurons Glutamate indicators with increased sensitivity and tailored deactivation...popular choice for a variety of synaptic imaging applications. Find iGluSnFR4 plasmids here! Aggarwal, A...., Iyer, R., … Podgorski, K. (2025). Glutamate indicators with increased sensitivity and tailored deactivation...DNA vector that contains everything needed to replicate inside a host cell but with space to insert your...

...resume that will give you a chance at an interview:  1) Have an excellent resume ready to be submitted The...demonstrate in your application). Beware the myths of scientist “overqualification” but it may not be ...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... here for more on timely submission of a job application. 3) Read job descriptions carefully Be open minded...bold. You don’t have to have all of the listed qualifications to apply for a position.  A job description...target someone in a specific department and/or location. The more people you meet the better chance you...

...and drawbacks of four split AAV vector approaches. 1. Overlapping  Overview: The overlapping strategy has...Adeno-Associated Virus Capsids. Human Gene Therapy Methods 27:1–12 . https://doi.org/10.1089/hgtb.2015.140 Frischmeyer...that the oversized AAV genomes are fragmented or truncated when packaged and then following transduction,...and may need to be optimized for each unique application (Trapani et al., 2015).  Expression of smaller...transgene to help prevent the accumulation of such truncated proteins. Examples of use:  Ghosh et al., 2008...design, donor A may be capable of expressing a truncated protein. This can be avoided by excluding a stop...JS (2011) Gene Therapy of mdx Mice With Large Truncated Dystrophins Generated by Recombination Using rAAV6...

...advocacy, and writing legislation (see Figure 1 for career map). The beautiful part of science policy is...about science policy as a profession. Figure 1: Science policy can lead you to many paths! For a ... policy implementation, program management, communications, diplomacy and international relations, advocacy...they train and bring researchers to Congress to advocate for increased science funding and other issues...sets for a science policy career is science communication so an outsized way to prepare is to do lots ...Initiative.    Get practical experience in science communication by writing for the Journal of Science Policy...connect you with alumni before and during your application process so you can learn more! Figure ...

...et al., 2019; McLaughlin, 2019).    Figure 1: Schematic of multiplex immunofluorescence approaches...). Springer New York. https://doi.org/10.1007/978-1-4939-8935-5_26 Jambor, H., Antonietti, A., Alicea,...protein is (localization) and how much there is (quantification). But what if you’ve got several interesting...primary antibodies that have been validated in IF applications (find IHC validated or ICC validated antibodies...informative image-based figures for scientific publications. PLOS Biology, 19(3), e3001161. https://doi....Immunocytochemistry Protocols for other antibody applications...

...organized and boost your productivity in 2014. Tip 1: Use color coding, labels, and post-its to make finding...moving your paper spreadsheets for reagent storage locations, such as the list used to organize your cryo freezer...

...information Since the inception of Phage Directory ~1 year ago, Sacher and Zheng found that they could expand...researcher may only know how to prepare phage for one application but not another or they may have only used phage...Directory expands, it can also fill this role by cataloging and archiving information on individual phages...

...Molecular and cellular endocrinology 215.1-2 (2004): 1-10. PubMed PMID: 15026169. Soo, Rochelle M., et al... into a pulsating display imitating a moving caterpillar. The worm controls the snail’s behavior, forcing...exposed to predators. Hungry birds spots the “caterpillar” and then poke out the snail’s eyes and ingests...

...strategies in the comments section!   References 1. Yearly Planning Meetings: Individualized Development...determining meaningful solutions. PI-trainee communication builds a positive lab culture Although they ... communication...

...Kan, SpHIS5, or CaURA3 selection marker. Lee et al.(1) assessed many of these fluorescent tags in S. cerevisiae...insert will typically contain both a protein modification sequence and a selectable gene product for isolation...tags have been integrated. Simply design your amplification primers with the desired targeting homology—...cerevisiae, looking at their performance in categories such as brightness, stability, and disruption of...optimal FP combinations for use in yeast imaging, categorized by specific filter sets and experimental output...This collection is ideal for tandem affinity purification of protein complexes. John Pringle and Jürg ...deposited a large collection of plasmids for genome modification in yeast developed by Dr. Pringle’s former lab...

...literature? Very often the data of interest are in 1 or 2 figures from one paper and then another figure...include: Cataloging a reagent’s performance as in this antibody ReFigure Analyzing replications as in this... ReFigure. Reading and exploration including replications and experiments resulting in “negative data”...small selection of data found in full-length publications. As early career researchers, we wondered how...reagent performs well, or you might publish a replication experiment that you conducted as part of setting...papers as they are not included in traditional publications. By submitting your data to a repository (Figshare...-registration, preprints, data, software, micropublications) in grant proposals. ReFigures are one such...

...datasets by computational biologists. Figure 1: The BioGRID Open Repository of CRISPR Screens (ORCS...   Figure 3: Each publication featured in BioGRID ORCS has a dedicated summary page with PubMed...favorite gene was described in a CRISPR screen publication. You see this mentioned in the results section...manually curated and annotated from more than 45 publications, beginning with the seminal research articles...screen curated in ORCS is linked to the original publication via its PubMed identifier, which can also be ...a list of individual screens curated for the publication. Clicking on the screen name takes users to a... supplemental data tables available with the publication are also provided for the user. This approach...

... everyone was very approachable.     Fig. 1: Examples of DNA sequencing QC work    In addition...working in your chosen field. This cooperative education experience (co-op) is what has brought me, a second... QC/QA team as a DNA Sequencing Intern. The application process When I first saw the comprehensive job...for the intended position. After submitting my application, I was contacted to participate in a brief fifteen...the depositing labs intended. With the use of applications like SnapGene and Geneious, I was able to determine...currently working on projects for the Science Communication function at Addgene. Working a 9-5 As it turned...

... if the name was the correct one!)    Fig. 1: HeLa cells. Image courtesy of Josef Reischig.  ...compounds, links to publication references, and cross-references with cell line catalogs and collections....surveying of cell line collections websites and catalogues Literature searches utilizing Google Scholar ...include accession number, species, and assigned category (cancerous, hybridoma, etc.), as well as sex and...manually going through the pdfs of the 26,000 publications cited. If the information seems incomplete or...

...social media? No problem - see our recap below! Day 1: gRNA design tools: How to Design your gRNA for CRISPR...job - we strive to also give you high-quality educational resources to help you do better research. CRISPR...design, sorting the plasmids we have available by category. Each plasmid page now has a helpful graphic to... newest page in the Transcriptional Regulation category already has five blue flame plasmids covering ...providing an avenue for temporary/reversible CRISPR modification. In addition to these subpages, our updated ... HypaCas9 Day 10: Transcriptional regulation: Truncated gRNAs for Regulating Gene Expression and CRISPR...

...of experiments they are commonly used for.  Table 1. Points to Keep in Mind When Performing Biosensor ..., and you should always check the associated publication for the specifics of your chosen plasmid. Types...conformational changes, translation, and protein translocation. Targeting your biosensor Before expressing ...Experiments Experiment  Detection Method Notes Quantification of a process Plate-based assays (plate reader...components of individual cells Separation/identification of fluorescently-labeled components Fluorescence-activated...

...plasmids. The Zhang lab, on the other hand, uses a 1 plasmid, single infection system. Genome wide mouse...  “This enables you to do customized genetic modification on a scale that was really not possible before...genome engineering with CRISPRs was limited to modification of a few loci at a time,” Wang said. “It was... lentiviral libraries and their demonstrated application offer proof-of-principle that CRISPR-Cas9 can...

...target genes, but we’ll save that application for another day! References 1. Tanenbaum, Marvin E., et al....aggregation, it’s recommended for most imaging applications. What can you do with SunTag? Tanenbaum et al...al. “A protein-tagging system for signal amplification in gene expression and fluorescence imaging.” Cell...

...that work for our experiments!)     Figure 1: The IBEX Imaging Community front page. That’s...something doesn’t work, it’s not a failure. It’s an education.   Figure 4: Zenodo page showing the IBEX... repository for reagents, protocols, panels, publications, software, and datasets. It’s also a great place...

...plasmids CRISPRi in Caulobacter crescentus Figure 1: Serial dilutions of CRISPRi in Caulobacter cells ...expression of specific Caulobacter genes using a catalytically dead Cas9 (dCas9) derived from Streptococcus...methylation-silenced genes. Read the paper in Nature Communications Find Casilio-ME plasmids If you have a new...

... successful postdoc fellowship. 1. Reverse engineer success On Day 1 of your postdoctoral experience,...need? Multiple first or corresponding author publications (published, not in preparation) A history of...The other items will follow. If you get more publications, you will be more competitive for fellowships...to write as well as mentor. Yet, even the most dedicated mentor is not always available for mentees due... project. Furthermore, Group Leader position applications require 3 letters of reference- the graduate...interests, complementary perspectives, a shared communication style. You “get” each other. The other factor...makes time for you, as needed, this is a strong indicator that this person cares about your science and ...

...Last updated on Oct 1, 2020 by Aliyah Weinstein. This post was contributed by guest bloggers, Wenning...process check out our Mouse Modeling blog posts: Part 1: Genetically Engineered Mice and Part 2: Breeding ...interested in using CRISPR to develop therapeutic applications that better human health.     References Cong...

... antibody recognizes and binds to.   Figure 1: Labeled diagram of an antibody including Fc, Fab,...45. https://doi.org/10.1016/B978-012198382-6/50026-1 Wang R, Xiang S, Feng Y, Srinivas S, Zhang Y, Lin ...tool that can be used for a variety of research applications. Within the immune system, antibodies are produced...antibodies to bind proteins is useful for research applications as well because they allow scientists to target...150 kDa, a nanobody is 12-15 kDa.   Research applications for antibodies You can use antibodies for experiments...Let’s dive in and take a look at some of these applications. Western blot Western blots are used to detect... by binding to cell surface receptors. These applications are typically used for investigating signaling...

...communities  Table 1: Examples of defining your audience for science communication activities  Many people... can and do consume science communication.  Formal education is education that is provided through a structured...underlying all communications efforts.  Scicomm definitions (as used by Addgene)    Science communication, or science...time engaging with science communication efforts.  Scientific communication or scientific writing is peer...to note that it is the actual communication itself, not the communication method, that determines if something...scientific or science communication. A Tweet is still considered scientific communication if the language is... engage or disengage at will.  Informal education is education that is provided, typically ad hoc, for...

... towards understanding this disease. Figure 1: Scientists are continuing to do essential research...funding and volunteer opportunities, protocols, publications, and more to accelerate the pace of research...

... Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...

...proteins, DNA, and RNA used in CRISPR.   Figure 1: Overview of the parts of CRISPR. The bacterial chromosome...simplifying the CRISPR-Cas9 system for laboratory applications. At Addgene, we sometimes refer to the programmable...

...depends on what you are looking to study. Figure 1: A schematic showing the differences in chemogenetics...beneficial for use in therapeutic or behavioral applications where drawn out effects are required. Targeted...

...the life scientist or bioengineer.   Figure 1: Map of a plasmid with its elements described below...Description Origin of Replication (ORI) DNA sequence which allows initiation of replication within a plasmid...point for PCR amplification or sequencing. Primers can be exploited for sequence verification of plasmids...important. Our “Plasmids 101” series designed to educate all levels of scientists and plasmid lovers - serves...plasmids are small circular pieces of DNA that replicate independently from the host's chromosomal DNA....virulence. All natural plasmids contain an origin of replication (which controls the host range and copy number...Minimally, lab-created plasmids have an origin of replication, selection marker, and cloning site. The ease...

...ScFvs, based on community demand. Figure 1: Antibody against PSD-95 from the Trimmer Lab. Information...addgene.org if you would like to contribute to the catalog. The Catalog Will Start With Neuroscience Antibodies ...performance in various applications are included in the full webpage. Educational Resources As part of...resource, which includes recombinant antibodies, educational materials, protocols, and shared data, on our...sequence information for the antibodies in our catalog, so that you will know exactly what you are working...community about how the antibodies work in various applications, enabling you to better plan your experiments... improving research infrastructure, from our educational materials to our repository of open samples. ...

...you do just this, probably in less than a minute (1). Green Listed is a new software tool used to design...found through https://wermelinglab.com/.   References 1. Panda, Sudeepta Kumar, et al. "Green listed—a CRISPR...the user. This approach is useful for several applications as will be discussed below. The software could...contain multiple gRNAs targeting all genes in the indicated species and were suggested to be the best gRNAs...

...them you can do today! 10 ways to share your science 1)   If you don’t already make it a habit to talk – ... Foundation Round Table. In any field, good communication skills help when it comes to finding a job. ...an argument for taking the time to hone your communication skills. So, where can a graduate student or ...opportunities than ever before for scientists to communicate about their work and their interest in science...your area or at your institution; the news or communications office is a good place to start. It’s not unusual...about science writing as a career, the Science Communication Program at the University of California, Santa... your research stories. Consider practicing communicating your research by guest blogging here, just leave...

...level of application not present in the online class.   Four ways to improve your next manuscript 1. Be consistent...of any scientist’s career. We use writing to communicate our findings and share them with the world. If...simply writing down your data doesn’t guarantee a publication. There is an art to composing manuscripts and...field of microbiology with multiple high-profile publications. My classmates consisted of graduate students...paper will be generated. After you complete a publication quality figure, produce the figure, its legend...

...and genome modifications to reporter vectors (e.g. pBV-Luc) and gene silencing (e.g. pLKO.1 - TRC cloning... (e.g. 6xHis containing pDEST-HisMBP). After purification the protein can be released from the resin and...types of yeast expression vectors. Consider your applications and experimental design. You can create a new...more traditional approach of an autonomously replicating vector that carries either a wild-type version...a variety of other types of vectors for most applications used to study gene function. From making transgenic...

...could be kept frozen after collection. Figure 1: Paul included a voltmeter on top of the battery box...under harsh conditions. One example application from the publication is the removal of unwanted cDNA derived...the van der Oost lab. Phylogenetic comparison indicates that like GeoCas9 and ThermoCas9, IgnaviCas9 is...a thermostable Cas9 be useful? It can enable applications where reagents must be stored at elevated temperatures...

...year and happy reading! Top 10 blog posts of 2015 1. CRISPR 101: Homology Directed Repair Addgenie Chari...overjoyed to provide you with these fantastic educational resources and will be sure to keep our eyes open...used to generate inversions, deletions, and translocations. It also provides researchers with many different...repository use Cas9 to cut DNA, Addgene Science Communications Intern Mary Gearing explains how Cas9 homolog...make it easier to make gene edits at new genomic locations. 6. CRISPR 101: Non-homologous End Joining So ...

...help you decide if a master’s might be for you. 1. Are you interested in master’s-level jobs? Ultimately...technical know-how, and helped develop my science communication skills. Ultimately, my degree prepared me to...technical or product support specialist,” “field application scientist,” or “scientific or technical writer...for PhD applicants since many will also accept applications from master’s candidates with experience.  Use...

...specific DNA sequence in only 1 min. Using these lentiviral plasmids, catalytically inactive Cas9 (dSpyCas9)...makes them more adaptable to a broad range of applications. Recently, the Spiess lab developed a set of... with specific genomic loci. He made use of catalytically inactive dCas9 by fusing it to an engineered...

... to just that particular assay.      Fig. 1: A native protein (left) is denatured, allowing the...Conformation changes usually fall into one of two categories: ones induced by the researcher and ones that...naturally, by binding to other proteins, changing locations, or even just as part of performing a specific...it a transmembrane protein, or is it know to translocate or fold in certain conditions? It can help to...

...evidence through education.  My only criticisms of what I saw are that we, as a society, need to: 1) use more...impartial. The best science happens when scientists have 1) the courage to make mistakes and 2) the integrity...also a science communicator, UBC Public Scholar, and a member of the Vancouver SciCATs. Follow her on ...researchers and supporters of science marched to advocate for scientist participation in government, evidence...promote help motivate you to spend a little time advocating for science and getting yourself out there for...April because devaluing science has alarming implications for our public policies—many of which impact...scientist. I’ve worked tirelessly to get the publications, grant funding, and teaching experience that...

...targeting MNase to antibody bound proteins (Figure 1). Importantly, since Protein A has weaker affinity...interactions for subsequent sheering of DNA by sonification. Though it helps capture weaker or more transient...change offers a significant improvement and simplification of the original technique, specifically in ...

...updated version of BRET called BRET2. Figure 1: TRUPATH measures heterotrimeric G protein dissociation...Kadowaki S, Haga T, Takaesu H, Mitaku S (2002) Identification of G protein-coupled receptor genes from the...biosensors Resources on Addgene.org Browse the catalog of fluorescent protein plasmids Find biosensors...

...can also rank the plasmids by abundance, scaled to 1, to create a “cumulative wealth distribution” curve...would be a good time to talk about the amplification and verification processes we use to ensure high-quality...your own experiments. Library amplification 101 Library amplification seems straightforward: E. coli ...transformed with the library DNA, they grow and replicate, and then their DNA is harvested, much like one...library has its own depositor transformation and amplification protocol, but the overall workflow is similar...ll need to remember when undertaking library amplification and sequencing. Maintain representation of the...electrocompetent Stbl4 cells when performing amplifications, and we use electroporation to ensure a good...