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Showing: 551 - 600 of 1018 results

  1. Choosing the B(right)est Fluorescent Protein: Photostability

    Type
    Blog Post
    ...dependence on excitation power is important since different fluorescence imaging strategies use widely varying..., TIRF or spinning disk confocal use entirely different excitation power regimes. Therefore, it is unpredictable...the best way to compare the photostability of different fluorescent proteins? Measuring photostability...excitation power. By repeating these measurements for different fluorescent proteins and by comparing how the ...measurements reported in the literature are performed in different ways. We describe a number of issues with some...

  2. Plasmids 101: The protein expression toolbox

    Type
    Blog Post
    ...protein expression journey! Resources and References References Nabet et al. The dTAG system for immediate...expression of essential proteins (when knockouts aren’t viable). siRNA or RNAi are small, double stranded...days. There’s also a CRISPR for that! CRISPR interference (CRISPRi) targets a catalytically dead Cas9 ..., and expression is tunable by titration with different amounts of tetracycline. Tetracycline regulation...

  3. Hot Plasmids - March 2020 - base editors, GEVI, MoClo, and optogenetics

    Type
    Blog Post
    ...1013 - 1014 vg/kg, a dosage comparable to those currently used in human gene therapy trials. They further...reproducibility as scientists can assemble parts from different labs and experimental systems in their own research...mutants, determined protein expression levels from different promoters, and inhibited gene expression using...C-terminus of Cre (CreC) and the N-terminus of Cre (CreN) are fused to the truncated Phytochrome B (PhyBΔ...eye, muscle and epithelial cells, as well as in different stages of development, ensuring the breadth and...

  4. Antibodies 101: Affinity Reagents

    Type
    Blog Post
    ...reduced-in-size options often have reduced binding strength, half-lives, and/or specificity as well. They’...bispecific, meaning that each scFv can target a different antigen; binding of each can be either simultaneous...convert anyone to downsize their affinity reagents, trendy as that may sound. Validated antibodies are still... a scientist looking for that perfect fit!  References and Resources Additional resources on the Addgene...AddgeneBrowse our recombinant antibody plasmid collection References Grumezescu, A. M. (2018). Drug targeting and ...

  5. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    ...Emily P. Bentley in October 2024. References and Resources References Demanèche, S., Bertolla, F., Buret...functions of natural plasmids. Created with BioRender.com. Resistance (R) plasmids: microbial defense... also causes outbreaks of food poisoning, the difference is virulence plasmids! This category includes...Mclellan, S. L., Willis, A. D., Comstock, L. E., & Eren, A. M. (2024). A cryptic plasmid is among the most...

  6. Progress Towards a PAM-Free CRISPR

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    Blog Post
    ...mammalian plasmid here and edit on! References and Resources References Kwart, D., Paquet, D., Teo, S. et...sequence constraints, or at least has looser or different requirements. Luckily, there are some available...necessarily more flexible than NGG; they’re often just different (e.g. NGAG, NGAN, etc.). Additionally, alternative...al). Additionally, SpG Cas9 with the same PAM preference of NG was isolated by the Kleinstiver Lab (read...

  7. Fluorescent Proteins 101: Introduction to FRET

    Type
    Blog Post
    ...or undetectable FRET, well-characterized FRET reference standards can be used to validate FRET measurements...sophisticated questions about cellular processes. Currently, FRET experiments can probe protein-protein interactions...Borghi 2016) to a selected protein. Overcoming current limitations to permit the use of multiple FRET ... diseases and even assist in drug discovery. References 1. Baird, Geoffrey S., et al. "Circular permutation...biomolecular structure and dynamics in live cells." Current opinion in structural biology 34 (2015): 52-59....

  8. Split-BioID: An Improved Method for Studying Protein-Protein Interactions

    Type
    Blog Post
    ...28032891. This is a different version of Split-BioID. BirA* is fragmented at a different site than in Schopp... This can artificially select against PPIs that aren't stable outside the native cell context. The Split-BioID...of the BirA* fusion matters! Schopp et al saw different levels of fusion protein expression and biotinylation...research? Plasmids are available from Addgene! References Schopp, I.M., Ramirez, C.C., Debeljak, J., Kreibich...

  9. Simple CRISPR-based Epigenetic Editing: dCas9-directed DNA Demethylation

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    Blog Post
    ...whenever you hear it (whether at a scientific conference or a karaoke bar), you probably want to sigh....methyltransferase 1 (DNMT1) to copy methylation marks from parental to daughter strands of DNA. When dCas9 is bound...nearly 100% demethylation without the confounds inherent to dCas9-TET, allowing one to study the causal... CRISPR-based tools for epigenetic editing.   References and resources:  Additional Resources on the Addgene...Addgene.org Read our CRISPR Guide to Epigenetics References Charlton J, Jung EJ, Mattei AL et al. (2020) ...

  10. Getting Started with Behavioral Neuroscience: Standardizing Experiments and Using Viral Vectors

    Type
    Blog Post
    ... can only carry around 4.5 kb. Different AAV serotypes have preferences for specific cell types. For example...end of an experiment series would behave very differently (distracted) from the ones tested at the beginning...Experiment Behavioral scoring systems (experimenter differences in behavior observation); experimenter blinding...viral service for their behavioral studies. References Stoica L, Ahmed SS, Gao G, Sena-Esteves M. Gene...

  11. Savvy Advocates Needed to Navigate a Scientific Enterprise in Flux

    Type
    Blog Post
    ... to address the unique needs of students from different departments across the university and alterations...track postdocs nationwide. On this level, FoR is currently compiling a database of institutional responses...implementation of new strategies designed to cope with current increases. These two rulings along with progress...Howell -Founder of Howell Communications Philip Brenner, PhD - Assistant Professor of Sociology. University...public policy. Follow David on Twitter @driglar.   References: 1. Alberts, Bruce, et al. "Rescuing US biomedical...

  12. The Materials Science of Optogenetics Experiments

    Type
    Blog Post
    ...Several protocols have been written outlining the different steps and materials required for each [1, 2]. ...discussed above, each opsin variant has slightly different activation/inactivation kinetics. For example,...Biology from the University of Michigan and he is currently a Postdoctoral Fellow in the lab of Mary Jeanne... or follow him on Twitter @derekpsimonphd.    References 1. Gutierrez DV, et al. Optogenetic control of...channelrhodopsin for activation of CNS neurons. Current protocols in neuroscience / editorial board, Jacqueline...

  13. Magnetic Control of Proteins: More than a Dream

    Type
    Blog Post
    ...Find the MagLOV plasmid here!   References and resources References Duret, G., Polali, S., Anderson,...just be an artifact and eliminate sources of interference, but the result persisted. (Even better, multiple...be developed from other fluorescent proteins.  Currently, the team is working to identify how MagLOV’s ... of many optogenetic tools. And, since MagLOV’s parent protein AsLOV2 has already been adapted into a ...

  14. Antibodies 101: Flow Compensation

    Type
    Blog Post
    ...However, if your flow cytometer reads them out on different lasers, you should not need to compensate between... Guide Chart.  Pro tip! Some antibodies go by different names depending on your flow cytometer’s setup...look at the other channels you’re using first. A different fluorophore channel may be affecting your populations...first before you apply compensation values to your current channels. A notable exception If you have a sample...CD8+ T cells and cancer immunotherapies. She is currently a research technician in Arlene Sharpe's lab at...

  15. Twenty Years of Addgene Sharing: CRISPR

    Type
    Blog Post
    ...previous Addgene CRISPR publications.  References and resources References Pyhtila, B., Kasowitz, S., Leeson...with CRISPR pooled libraries, although we used different categories (Figure 2C). Note that in Figure 2,...through the years.  Libraries are divided into different categories based on function: knockout (KO), activate...have CRISPR plasmids deposited from labs in ~40 different countries, and we’ve shipped CRISPR plasmids to...

  16. Cloning Mammalian Cells with the Agarose Method

    Type
    Blog Post
    ... splitting transfected cells into 3 plates at different densities for selection). A 2% agarose solution...to heat- the inside of the wrist or the cheek. (Parents of infants will have no problem with this!) Make...transfection into plates containing selection agent at different densities, then dispose of the plates that do ...freeze at least 2 ampules each of 4 clones with different levels of expression of our gene of interest. ...to rapidly isolate individual cell clones with different expression characteristics from a population of...

  17. Fluorescent Proteins 101: Visualizing Subcellular Structures & Organelles

    Type
    Blog Post
    ... the protein within a living cell. Analyzing differences in protein localization and transport between...signals not visible to the human eye and are indifferent to subjective color intensity perception. Open...statistics that can be used to compare between different experimental setups. A colocalization analysis...one fluorescent signal in a channel setup for a different fluorescent signal) and photobleaching i.e. FP...Markers of Subcellular Localization Poster here! References 1. Allen Institute Plasmid Pages 2. Cooper GM...

  18. Of Myc and Men

    Type
    Blog Post
    ...antibodies, context is key.    References and Resources References Evan GI, Lewis GK, Ramsay G, Bishop...cell proliferation and tumorigenesis.  Using two different synthetic peptides corresponding to linear sequences... or the C-terminus, Evan, et al. isolated six different monoclonal antibodies (Evan, et al. 1985). Three...-9E10 variable regions and sticking them onto different scaffolds. But over the years, several alternative...

  19. No Llamas Required - Synthetic Nanobodies Against Membrane Proteins

    Type
    Blog Post
    ...randomized with a defined set of different trinucleotides (corresponding to different amino acids) in the regions...this, they injected the spike protein from two different coronaviruses into a llama over several days and...your target protein. The Seeger lab designed 3 different mRNA sybody libraries for use in the protocol,.... Find the Seeger lab's SARS-CoV-2 sybodies! References Geertsma ER (2013) FX Cloning: A Versatile High-Throughput...

  20. Golden Gate Assembly Upgrades: More Fragments, Faster Assembly, and Higher Fidelity

    Type
    Blog Post
    ...pathway engineering and optimization. However, current best practices for assemblies of more than 10 modules...rely on two-step hierarchical approaches using different Type IIS restriction enzyme specificities at each...Assembly test systems Our research focused on three different levels of assembly: High efficiency and accurate...products to market, New England Biolabs, Inc.       References 1. Vladimir, Potapov, et al. "Optimization of...cloning protocols Check out our molecular biology reference pages ...

  21. Plasmids 101: 5 factors to help you choose the right cloning method

    Type
    Blog Post
    ...still love using them! With so many technologies currently available, you now face the challenge of trying...create customized overhangs to assemble up to 6 different fragments in a prescribed order. These fragments...protocol is fairly flexible and adaptable to cloning different types of DNA, such as cDNA, PCR fragments, or ...deposit your finished constructs at Addgene!). References 1. Li, Chaokun, et al. "FastCloning: a highly...Find vector backbones Read our molecular biology reference ...

  22. Experimenting with New Careers while in Grad School

    Type
    Blog Post
    ... of "The world’s biggest and best children’s museum", The Children's Museum of Indianapolis. I drove passed...science outreach: engaging the public to promote an awareness and understanding of science. Like most science...advantage of my graduate experience. Melanie Fox is currently the founder and executive director of Central ...Plasmids for Your Research Learn About A Variety of Different Fields on Our Collections Pages  ...

  23. MXS Chaining

    Type
    Blog Post
    ... a nutshell, is the systematic combination of different genetic sequences into plasmid DNA. In high throughput...cloning can be used to combine the functions of different genetic elements to generate non-natural tools...assembling these modular types of constructs, and different methods are suited for particular downstream applications...Cloning Strategies Method Technology Pros Cons References Golden Gate Type II restriction enzymes Can...Kits at Addgene Find Plasmid Cloning Protocols References 1. Anderson JC, Dueber JE, Leguia M, Wu GC, Goler...

  24. Course-Based Undergraduate Research Experiences (CUREs)

    Type
    Blog Post
    ... in and out of the classroom.   References and Resources References Bangera, G., Brownell, S. 2014. Course-based...1977; Ma et al., 2020). Many have interpreted this trend as evidence that the best teachers and mentors pass...experiences afforded to those fortunate enough to apprentice with top scientists to larger groups of trainees...the University of San Diego), decided to take a different approach. His own most impactful learning experiences...

  25. The Effect of COVID-19 on Liu Lab Plasmid Requests From Addgene Data

    Type
    Blog Post
    ...capture the fact that different parts of the world encountered COVID-19 at different times. Most notably...papers. Figure 1: Plasmid requests by date. Parentheses depict the total number of plasmids ordered for...regions. Each of these regions was affected at a different time by COVID-19, so we defined the COVID-19 event...COVID-19 on many other scientific fields could be different.  Conclusion Figure 7: Mean inferred effects...

  26. DNA Purification Without a Kit

    Type
    Blog Post
    ...maxi, giga) which requires you to buy different kits for different purposes, with the Kim and Morrison ...Read our Molecular Biology Reference Visit our protocol video library References Kim, Y., & Morrison, S....proteins in solution. The pH is then lowered using a renaturing solution, which causes the proteins and genomic...

  27. CRISPR Between the Genes: How to Experiment with Enhancers and Epigenomics

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    Blog Post
    ...As with GPS in the real world, coordinates alone aren’t very useful. We’ll need something like Google Maps...denotes intersection and the – operator denotes set difference. That’s the intuition for how genome math helps...enhancers from step 2. See which ESCs die or differentiate Step 3 reveals which NANOG-related genes are...distinct enhancer disrupted. As our ESCs die and differentiate over time, we periodically use next-generation...by starting with all of the DHS sites from 125 different human cell types from the ENCODE Project. DHS ...

  28. Sharing Your Lab Protocols: Using Apps to Save Time & Track Your Experiments

    Type
    Blog Post
    ...your favorite protocols for future reference. The exchange currently boasts over 2,500 protocols covering...protocols.io/ Protocols.io is the creation of ZappyLabs. Currently in beta testing, the software aims to be a life...for others to use, and add your own comments. Currently you can find protocols from NEB, OpenWetWare (...101 articles on various plasmid features, and reference tables (i.e. antibiotic concentrations, commonly...

  29. Antibiotic Resistance: An Old Solution but a New Problem

    Type
    Blog Post
    ...multi-drug resistance are resistant to three or more different classes, or types, of drugs used to treat them...resistance can arise, and has arisen, for many different reasons. Exposure to antibacterial drugs provides...drug to work… if the instructions for treatment aren’t followed closely, you could be doing more harm ...bacteria are resistant to antibiotics and which ones aren't?     This video shows how to perform a Kirby-Bauer...science communication, and public health.     References 1. O'Neill, J. "Tackling drug-resistant infections...

  30. Viral Vectors 101: Preparing Pooled Libraries

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    Blog Post
    ...Addgenie Hannah Dotson.  Resources and references References Genome-scale CRISPR-Cas9 knockout and transcriptional...and producing lentivirus. But pre-made libraries aren’t quite plug-and-play: you’ll still need to titer...incubation time, antibiotic conditions, and amount of polybrene. (psst…didn’t get your library pre-made from Addgene...Addgene’s pooled libraries use these protocols.  References McDade JR, Waxmonsky NC, Swanson LE, Fan M. ...

  31. CRISPR 101: Which Cas9 Do I Choose for My CRISPR Experiment?

    Type
    Blog Post
    ...options in our CRISPR 101 series!  References and resources References  Anzalone, A. V., Koblan, L. W. &...plasmids that have been optimized for expression in different species and cell types including (but not limited.... Nat. Biotechnol. 2020 387 38, 824–844 (2020). Moreno-Mateos, M. A. et al. CRISPR-Cpf1 mediates efficient...101 Series Learn about the PAM Requirements of Different Cas9 variants Learn about CRISPR Pooled Library...

  32. Hot Plasmids - August 2020

    Type
    Blog Post
    ...For labeling the plasma membrane, they tested different target sequences for improved selectivity and ...into the supernatant. Four plasmids containing different nanobodies (NB) were generated in pMAK:  pMAK461...pluripotent stem cells By Jennifer Tsang CRISPR gene interference (CRISPRi) and gene activation (CRISPRa) are ...Addgene! A new dual-deaminase base editor enables concurrent adenine and cytosine editing. Find the plasmids...oscillator, bistable network, stripe pattern-forming incoherent feed-forward loop. Find the plasmids at Addgene...

  33. Supporting Reproducibility with a Connected ELN

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    Blog Post
    ...critically important to be able to conveniently reference specific pieces of data stored externally in the...of research compiled in the ELN. Without such references the record of research is incomplete, making ...research Just as researchers need to access and reference external data sources, they also need to continue...spreadsheets, images, sequencing data, etc.) used and/or referenced in the experiment. Since most ELNs, unlike data...produced, collected, and analyzed. While the many different ELNs out there can help with searchability, record-keeping...

  34. Data Freedom: The Expansion of Data Sharing in Research Publications

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    Blog Post
    ...of veracity of the conclusions drawn. The main difference with the PLOS policy is that access of this data...perhaps similarly impoverished lab) might apply different or novel tools and assumptions to reveal findings...geneticists may track the motion of multiple animals on different occasions via video, generating huge amounts of...albeit unintentional, tendency to work against transparency. This opens science to charges of elitism and...opportunities for research as well as increasing transparency. Thank you to our guest blogger! Jim Woodgett...

  36. Adeno-associated Viruses (AAVs) for Genome Editing

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    Blog Post
    ...comparable or better than the efficiency of most current CRISPR-based approaches for the generation of knockins...applications that require large gene fusions. The current state-of-the-art in combining the AAV and CRISPR-based..., Georgetown School of Medicine. His research currently focuses on determining the mechanisms and consequences...cohesin gene inactivation in human cancer.     References 1. Russell, David W., and Roll K. Hirata. "Human...vectorization of DSB-mediated gene editing technologies." Current gene therapy 16.3 (2016): 207-219. PubMed PMID:...

  37. CRISPR 101: Validating Your Genome Edit

    Type
    Blog Post
    ...dice on a new gRNA in the future! References and Resources References Eva K. Brinkman, Tao Chen, Mario ...your bulk population. This way you can ensure you aren’t looking for a needle in a haystack before going...option, especially if the options outlined above aren’t realistic. Check Out Our Post on Sequencing Options...M, Schönig K, Eckert H, Eschstruth A, Mianné J, Renaud J-B, Schneider-Maunoury S, Shkumatava A, Teboul...

  38. Quick Guide to Near-Infrared Fluorescent Proteins

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    Blog Post
    ...recent research @daria_gift_shch.     References and resources References Shemetov, A.A., Oliinyk, O.S. & ...phytochrome (BphPs) photoreceptors have higher apparent cellular brightness than previously reported widely...the monomeric state of a probe leads to less interference with the oligomeric state of a fusion partner...Proteins: Multiplexing and Optogenetics across Scales. Trends Biotechnol 36, 1230-1243 (2018). Shcherbakova, ...

  39. Antibodies 101: Single Chain Fragment Variables (scFvs)

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    Blog Post
    ...delivering scFvs that inhibit HIV.   References and Resources References: Ahmad ZA, Yeap SK, Ali AM, Ho WY...single chain fragment variables (scFvs)? scFvs weren’t developed for another decade when in 1988 they ...additional rounds of screening. Created with Biorender.com.   Some phage display libraries screen antibody... the prevention or treatment of HIV infection. Current Opinion in HIV and AIDS 10:190–197 . https://doi.org...

  40. Fluorescent Proteins 101: When GFP lets you down

    Type
    Blog Post
    ...purified protein at different acid concentrations. Overviews of the pKa values of different fluorescent protein...requires an N-terminal consensus sequence (MG-) and prenylation requires a C-terminal consensus sequence (-CaaX...can follow him on twitter: @joachimgoedhart.   References 1. Zacharias, David A., et al. "Partitioning ...LOV-based reporters for fluorescence imaging." Current opinion in chemical biology 27 (2015): 39-45. PubMed...

  41. CRISPR 101: Engineering the Plant Genome Using CRISPR/Cas9

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    Blog Post
    ...in a wide variety of organisms.  While much of current CRISPR research focuses on its potential applications... region into the plant genome. Created with BioRender.com. In recent years, deaminase-mediated base...agrobacterium-mediated delivery. Summary While different in many of the specifics - promoters used, precise...mediated genome engineering in plants isn’t all that different from how it’s used in other systems. Luckily, ... don’t be afraid to try your hand at CRISPR. References Belhaj K, Chaparro-Garcia A, Kamoun S, Nekrasov...

  42. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

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    Blog Post
    ...Drosophila. Why Brainbow? Brainbow highlights the trend in biology towards single-cell resolution imaging...bottom of this post for a basic synopsis of the differences amongst the Brainbow systems. Pre-Brainbow techniques...controlled by FLP/FRT recombinase. When placed under different promoters, Brainbow and Flpbow can be used to ...strategy described in Karine Loulier, et al. (2014). Currently, Brainbow techniques are also being applied to...2.1 plasmids Brainbow 3.0, 3.1, 3.2 plasmids References: Transgenic strategies for combinatorial expression...

  43. A Control for All Seasons

    Type
    Blog Post
    ...check out these blog posts.       References and resources References Burry, Richard W. “Controls for ..., you will have a better understanding of the different types of controls and be able to use this information...treatment versus and which are likely just natural differences between individuals. Recommended controls for....  Western blotting When comparing samples in different lanes in a western blot, you’ll need to include...

  44. Recombinase-based State Machines Enable Order-dependent Logic in vivo

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    Blog Post
    ...register (Fig. 1C below). If attB-attP pairs from different recombinases are overlapping or nested on a register...that enters distinct states when treated with different combinations and orders of two recombinases. ...or strings of parts may be PCR-ed out of their current registers and used to build new registers. The ... register. The purpose of this is to prevent interference between the transcribed gene and its 5’ UTR.... in biocomputing and biological circuits.     References 1. N. Roquet et al., Synthetic recombinase-based...

  45. Plasmids 101: Restriction Cloning

    Type
    Blog Post
    ...their different migration speeds; after running the gel for some duration of time, these differently sized...time required for complete digestion varies for different enzymes. Many companies now sell fast digest enzymes...purification. In gel purification, you use a voltage difference across a gel matrix (usually agarose) to pull...sized pieces will be in different locations and can be cut out of the gel individually. There are a variety...

  46. Activate Addgene's "Find Plasmids" Badge on Pubmed Abstracts

    Type
    Blog Post
    ...“NCBI Site Preferences” box. Once on the Preferences page, navigate to “PubMed Preferences” and click ... that you know how to navigate to the PubMed Preferences and Filters pages, be sure to check out the other...

  47. Streaking for Single Colonies: The Streak Plate Challenge

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    Blog Post
    ...He mixed together 10 E. coli strains producing different pigments, and tasked the students to separate ...over 20 years. Previously, the lab used three different species for the challenge, but one species always...Addgene is have the students transform them into different strains of E. coli,” says Westenberg. During the...transformation experiment, he plans to show how different genetic backgrounds of the host strains affect...

  48. Visualizing Your Plasmid Request Data

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    Blog Post
    ...requested plasmid, the number of requested plasmids, or trends of requests over time. Visualizing the data So,...is a great step up, but I needed a way to show different aspects of the data simultaneously. That’s why...dashboards are (i) easy access to important numbers and trends, (ii) user interaction with the data, and (iii)...new data. These features are incorporated in the current dashboard. The dashboard is coded in R Markdown...

  49. Lambda Red: A Homologous Recombination-based Technique for Genetic Engineering

    Type
    Blog Post
    ...allele is selectable, it can be transferred to a different genetic background via P1 transduction. Plasmid...could be used to move the modification into a different background, if needed. This approach is best suited...our guest blogger, Beth Kenkel Beth Kenkel is currently a research scientist in the Department of Laboratory...science communication and in vitro diagnostics.     References 1. Costantino N, Court DL. Enhanced levels of...Engineering in Bacteria Using Homologous Recombination. Current Protocols in Molecular Biology. 106:V:1.16:1.16.1...

  50. Microbes: Look, Listen, and Tell

    Type
    Blog Post
    ...When you think about going to a scientific conference, you may think about sitting amongst a sea of chairs...ultrasound dyes because these dyes “collapse” at different ultrasound frequencies. Read more about ‘Using...created that produces loads of inspiration for the current and the next generation of microbiologists. If...communication check out these SciComm blog posts. References 1. Bourdeau, Raymond W., et al. "Acoustic reporter...
Showing: 551 - 600 of 1018 results