...joined Sanofi/Genzyme nearly three years ago I decided it was time to get in shape. After three kids, ... to raising funds and awareness of FOP. Please consider running with us or making a donation via the link... who is a big superhero fan. This year, I have decided to run as The Green Lantern’s friend, St Walker...

... how microbiology research is improving lives worldwide and the presentation made it clear how important...up with former colleagues! I ran into past ASM President Tim Donohue (see below) on my way back from getting... with my favorite microorganism (Rhodobacter sphaeroides!) at the ASM booth. As well as being the Director...

...reporters that are detectable by an inexpensive and widely available imaging platform: ultrasound. Challenges...experiments because  ultrasound devices are inexpensive, widely available, and have deep tissue penetration and... imaging. Specifically, arg1 expressing E. coli aided in ultrasound imaging of the mouse gastrointestinal...

...viewing sequencing traces, while the paid version provides more tools for plasmid mapping and design (Figure... deposited with Addgene. NCBI BLAST: Compares nucleotide or protein sequences to sequences in a database...molecular biology protocols Browse our protocol video collection ...

...know that the Fc domain provides structure, determines isotypes, and provides a place for secondary antibodies...yes, it does! While the Fc domain’s functions, besides structure and isotype determination, don’t generally...

...conversations. This creates a level of familiarity and confidentiality to ensure more honest answers. Ask people who...resources to help you choose a good mentor, I have provided a downloadable PDF at the end of this blog post... may hedge on their answers to your questions. Consider reading Toxic Academic Mentors by @drmellivora...

...methods for data analysis, our team’s goal is to provide scientists with new tools and protocols that they...labs looking to improve their AAV yields. Given the wide range of downstream applications scientists rely...resources of the Addgene blog Read our beginner's guide to viral vectors Find all of our viral vector blog...

....  These days commercially available TOPO kits provide vectors or cloning arms with overhanging 3´ deoxythymidine... 2. Novel approach to molecular cloning and polynucleotide synthesis using vaccinia DNA topoisomerase....Molecular Biology Techniques Watch Our Protocol Videos ...

...biological tools, and with the right safety considerations, they aren't dangerous for controlled lab usage...viruses later on. These built-in safety features provide extra layers of risk management when working with...short-term? Answering these questions will help guide you to the right method for your experiment.  A ...

...of inquiry with which I was not yet familiar. The idea that my piecemeal degree might one day foster a ... several projects nearing deadline (an animated video depicting nuclear fission; a podcast with a genetics... appeal to a given audience and how to utilize a wide array of media in doing so—can prove invaluable ...

...cellular-level events and behavior. It’s a problem that he considers “one of the greatest challenges of neuroscience...particular advantage for observing events taking place inside cells that are on the move, such as T lymphocytes...protein, troponin C (TnC), from muscle cells. They identified a toadfish TnC variant which had a high-affinity...

...brief history of optogenetics? How long has this idea been around? Boyden: Optogenetics is a toolset that... have tried to make green molecules, but then alongside blue they actually have a lot of crosstalk. So...for the first time? Boyden: There are always considerations for each species. This could be a three or ...

...that interact with the virus. As a result, we identified 332 human proteins that the virus interacts with...the help of human proteins in their lifecycle, identifying these potential human helpers of SARS-CoV-2 (...You could be the next scientist joining in this worldwide fight against SARS-CoV-2. As we’ve all been affected...

...center around a cast of endearing characters who decide to take matters into their own hands. Who doesn..., I wanted to be challenged to read some books outside of my comfort zone. I’d been in a bit of a reading.... We’ve read well-known classics, like Austen’s Pride and Prejudice, and some more obscure treasures, ...

...gene expression in a target cell. Practical considerations If you have high quality AAV, you can address... desired titer and want to dilute it, err on the side of caution when adding buffer back to the solution... for Work with Virus Watch Our AAV Purification Video ...

... 3 hours. It was robust, capable of generating a wide range of insert sizes and also produced high yields...interact and learn more about one another. They also provided both breakfast, lunch, and snacks. At the end ... to constantly update our processes and get new ideas from learning experiences like workshops at Lab ...

... Science. For each publication, Web of Science provides basic bibliographic information and uses its vast...is very plausible that science published in more widely-read journals gets more citations and is more likely...citations due to the journal of publication. Table 1 provides an example calculation of the weights. Table 1...

...contrast” variant provides improved detection for tracking large populations of neurons in wide-field imaging...Addgene's viral service Find viral protocols and videos Find AAVs for your research ...

...is currently using VAMP-seq to study vitamin K epoxide reductase (VKOR) variants. VKOR is an enzyme critical...well-known but poorly studied neuromodulator, and evidence suggests that it is incredibly important for nearly...use the serotonin receptor to study how single nucleotide polymorphisms present in the human population...

...model yeast strain and discuss why you may want to consider a yeast off the beaten path.   Yeast as a model...disease models established in yeast, including a widely used Parkinson’s experimental model. In fact, approximately...of K. phaffii lacks certain terminal mannose glycopeptides (which most other yeast possess) that are problematic...

...used to infer differentiation trajectories and identify both up- and down-regulated pathways. Individual...unique barcodes for each TF help streamline identification. While the Zhang lab controlled their multiplicity... a model system. TF screens can be designed to identify custom TF programs to acutely initiate disease...

...CRISPR locus of a bacteria host encodes multiple guide RNAs (gRNAs) on a single array to target the genome...applications when repetitive tRNA sequences must be avoided, different tRNAs with optimized 5’ leader sequences...array for multiplex gene editing applications, we provide detailed manual and online design examples of using...

...coinjection into zygotes of Cas9 mRNA and a single-guide RNA targeting the mutant allele (7). An independent...CRISPR/Cas9 has been used to generate mutations in a wide range of organisms, but its potential to efficiently...These papers suggest it might work well enough to consider as a therapeutic.” There are many fundamental ...

...the pMinDisplay backbone is fused to a specific peptide segment (PDGFR TMD) that anchors the sensor to ...the membrane and displays it on the extracellular side. In addition to the original PDGFR TMD, the researchers...ordered directly from Addgene. The Podgorski Lab has provided a brief breakdown of the promoter systems you ...

...bioscience and tech transfer communities who help guide and promote Addgene in its mission to make research...given by Phil Sharp. After a few rotations, Doench decided to complete his PhD work in Sharp’s lab.  Doench...a tremendous amount of technology development alongside it.” Doench starting working with David Root ...

...proteins to be imaged together, you will need to consider the same factors as when choosing an individual... the filter sets or lasers on your microscope. Ideally, you would like to have substantial overlap between...protein and a filter set, many filter set vendors provide tools to plot the fluorescence spectra of proteins...

...detection of the reporter in a specific assay, which ideally would have low background signal, high sensitivity...bioluminescent reporter, other luciferases have been identified in various species with differing kinetics, substrate...luciferase used, a compatible substrate must be provided for the enzyme to catalyze the light-producing...

...based on maps of protein structure. Sometimes this provides a great deal of clarity. But, often these methods...Institute Target Accelerator Plasmid Collections, now widely available to the scientific community through Addgene...Functional Interrogation of Rare Cancer Variants Identifies Oncogenic Alleles. Kim E, Ilic N, Shrestha Y....

...check out this blog post or see Addgene's CRISPR guide page for a review of Cas9.  For a brief comparison...Essletzbichler, P.,  Zhang, F. (2015). Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. Cell...Research Catch up on Your CRISPR Background with Our Guide Pages ...

...there are many great resources available to help guide you through the rough waters of mutation validation...great deal of precision. This means digital PCR provides an absolute measure of target abundance, does ...screening to validate your genome edits, this paper provides detailed instructions on how to design such assays...

... and gotten unusual or unexpected results? Or considered why DpnI will degrade your template DNA from ...methylated (if it comes from a phage for instance). Aside from restriction modification systems, DNA methylation...level, the potential for methylation should be considered when digesting DNA. Why? Well, even though Dam...

...task and will prevent you from getting lost in the wide word of informatics. I am a molecular biologist ... project a few years ago when my advisor and I decided to do a massive RNA-seq analysis. I had never interacted...even if your conclusion is “this approach is not ideal because…” Including a conclusion for each analysis...

...through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ... the first reporter genes using ultrasound and provide a major benefit to scientists studying microbiome...different cells present in a mixed population, or identify differentiation and disease states. This collection...

...promoter should flank this component as well. On both sides of this promoter-gene combo are two long terminal... and have appropriate factors displayed on the outside of the particle to interact with the host. This...applications and aid in ease of use. Our lentiviral guide outlines these components.     Fig. 1 - ...

...within a biological system. Basically, biosensors provide quantifiable readouts for levels of different chemicals...FRET-based biosensors have been engineered to detect a wide array of neurotransmitters ranging from dopamine...fluorescence-based and have been optimized for a wide-range of uses to monitor the fundamental cellular...

... kB, then you would be oversized even before considering the need for some manner of polyA. In this scenario...can be safely concentrated to ~1x1014 vg/ml.. As evident in our repository, this application is an outlier...Addgene.org Find Adenovirus Plasmids Adenovirus Guide Page Subscribe to Viral Vectors Blog Posts ...

...through our hot plasmids articles. These articles provide brief summaries of recent plasmid deposits and ...can be reconstituted in vivo to form a protein identical in sequence to the unmodified enzyme. Each half...any exogenous sequences and regenerates a native peptide bond at the split site. Figure 1: The base...

...interactome of these key regulatory molecules, and has provided clues to their previously uncharacterized cellular...acetyltransferase to luciferase, the basic assay remains widely used today.  The sheer abundance of these critical...greatly increased the burden associated with identifying plasmids in the research literature and acquiring...

...specificity of AAV, work has focused on modifying the outside protein shell, or capsid, of the virus so that ...here.  Applications for synthetic promoter AAVs Besides restricting AAV transgene expression to particular...synthetic promoters with high rates of labeling are ideal of gene therapy applications, promoters that produce...

...initiation step of gene transcription; today we'll provide an overview on how transcription stops, or termination...post-transcriptional additional of multiple adenine (A) nucleotides to the tail of a messenger RNA transcript. The...be detailed here, but additional references are provided at the end.  Nearly all common bacterial expression...

...molecular tools to map neuronal connectivity are widely used in neuroscience. In this post, I’ll describe... in first order neurons, the G-deleted RABV can divide and replicate (since these processes are mediated...Another study used RABV monosynaptic tracing to identify inputs to two major dopaminergic cell populations...

...Even before fluorescent proteins (FPs) came into wide use, there were a variety of ways to monitor cell...few properties that have now become hallmarks of ideal FP timers: 1. Ratiometric determination of expression...Empty Backbones Check Out Our Fluorescent Protein Guide ...

...to not produce artifacts in your experiments. Considerable effort has gone into making FPs monomeric, yet..., increasing the probability that two FPs will collide with each other. The consequences can be dramatic...localized to the endoplasmic reticulum, the normally spiderweb-like pattern of tubules can be grossly distorted...

...SpCas9. Hu et al. performed GUIDE-seq analysis to characterize genome-wide off-targets, since increasing... Resources on Addgene.org Check out our CRISPR Guide Find CRISPR Plasmids for Mammalian Systems Learn...

...that popularity through the years.  Libraries are divided into different categories based on function: knockout... years, even as more CRISPR technologies become widely used and available. If we look closely at Figure...navigate that learning curve. As of 2023, our CRISPR guide has been viewed 1,904,462 times, and the CRISPR ...

...Degrons are destabilizing domains which will be identified by various enzymes and sent to the proteasome...protein for imaging. Given its popularity and widespread usage, a proteolysis-targeting chimera (PROTAC...some stage, then choosing a HaloTag could pay dividends down the road. It is difficult to predict precisely...

...below. Types of Voltage Indicators GEVIs can be divided up structurally into 2 main groups: voltage sensitive...voltage sensing component (i.e., rhodopsin naturally resides in the membrane), or both. Targeting these systems...Addgene’s Virus Protocols Addgene’s Chemogenetics Guide Resources on the Addgene blog Viral Vectors ...

...factor of 2. This means that if you set up two identical assays with the same sample, you could end up ... should fall within the range of the assay for a wide range of titers. We usually dilute our samples 1...experiments.  References Furuta-Hanawa, Birei, Teruhide Yamaguchi, and Eriko Uchida. "2D droplet digital...

...sites. FRT sites and loxP sites differ at the nucleotide level but share an overall structure of 13 base...37 °C). Smart molecular evolution led to the identification of FLPe which has a temperature optimum of ...Recombination between the two target sites with identical sequence leads to inversion of the flanked DNA...

... codon, stop codons, reading frame considerations, etc). This video demonstrates how to use the Snapgene...Protocols Watch Our Diagnostic Restriction Digest Video Check Out Our Molecular Biology Reference Pages  ...